| 1. |
- Tapani, Sofia, 1982
(författare)
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Stochastic modelling and analysis of early mouse development
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The aim of this thesis is to model and describe dynamical events for biological cells using statistical and mathematical tools. The thesis includes five papers that all relate to stochastic modelling of cells. In order to understand the development and patterning of the early mammalian embryo, stochastic modelling has become a more important tool than ever. It allows for studying the processes that mediate the transition from pluripotency of the embryonic cells to their differentiation. It is still unclear whether the positions of cells determine their future fates. One alternative possibility is that cells are pre-specified at random positions and then sort according to a already set fate. Mouse embryonic cells are thought to be equivalent in their developmental properties until approaching the eight-cell stage. Some biological studies show, in comparison, that patterning can be present already at sperm entry and in the pronuclei migration. We investigate in Paper I the dynamics of the pronuclei migration by analysing their trajectories and find that not only do the pronuclei follow a noise corrupted path towards the centre of the egg but they also have some attraction to each other which affects their dynamics. Continuing in Paper II and III, we use these results to model this behaviour with a coupled stochastic differential equation model. This enables us to simulate distributions that describe the meeting plane between pronuclei which in turn can be related to the orientation of the first cleavage of the egg. Our results show that adding randomness in sperm entry point is different from the randomness added through the environment of the egg. We are also able to show that data sets with normal eggs and eggs treated with an actin growth inhibitor give rise to considerably different model dynamics, suggesting that the treatment is affecting the migration in an invasive way. Altering the pronuclei dynamics can alter the polarity of the egg and may transfer into the later axis-formation process. Invasiveness of experimental procedures is a difficult issue to handle. The alternative to invasive procedures is not appealing since it means that important developmental features may not be discovered because of individual variability and noise, leading to guesswork of the underlying mechanisms. The embryonic cells are easily affected by treatments performed to make the measuring, made by hand, easier or by the light exposure of the microscope. Treatments as such are used for example for producing flourescent proteins in membranes or slowing processes down. Paper IV and Paper V serve to analyse how light induced stress affects yeast cells and we employ a method for analysing the noisy non-stationary time series, which are a result of the yeast experiments, using wavelet decomposition.
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| 2. |
- Wieloch, Thomas, 1979-, et al.
(författare)
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Intramolecular carbon isotope signals reflect metabolite allocation in plants
- 2022
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Ingår i: Journal of Experimental Botany. - : Oxford University Press. - 0022-0957 .- 1460-2431. ; 73:8, s. 2558-2575
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Stable isotopes at natural abundance are key tools to study physiological processes occurring outside the temporal scope of manipulation and monitoring experiments. Whole-molecule carbon isotope ratios (13C/12C) enable assessments of plant carbon uptake yet conceal information about carbon allocation. Here, we identify an intramolecular 13C/12C signal at tree-ring glucose C-5 and C-6 and develop experimentally testable theories on its origin. More specifically, we assess the potential of processes within C3 metabolism for signal introduction based (inter alia) on constraints on signal propagation posed by metabolic networks. We propose that the intramolecular signal reports carbon allocation into major metabolic pathways in actively photosynthesizing leaf cells including the anaplerotic, shikimate, and non-mevalonate pathway. We support our theoretical framework by linking it to previously reported whole-molecule 13C/12C increases in cellulose of ozone-treated Betula pendula and a highly significant relationship between the intramolecular signal and tropospheric ozone concentration. Our theory postulates a pronounced preference for leaf cytosolic triose-phosphate isomerase to catalyse the forward reaction in vivo (dihydroxyacetone phosphate to glyceraldehyde 3-phosphate). In conclusion, intramolecular 13C/12C analysis resolves information about carbon uptake and allocation enabling more comprehensive assessments of carbon metabolism than whole-molecule 13C/12C analysis.
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| 3. |
- Blockhuys, Stephanie, 1983, et al.
(författare)
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Defining the human copper proteome and analysis of its expression variation in cancers.
- 2017
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Ingår i: Metallomics. - : Oxford University Press (OUP). - 1756-5901 .- 1756-591X. ; 9:2, s. 112-123
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Tidskriftsartikel (refereegranskat)abstract
- Copper (Cu) is essential for living organisms, and acts as a cofactor in many metabolic enzymes. To avoid the toxicity of free Cu, organisms have specific transport systems that 'chaperone' the metal to targets. Cancer progression is associated with increased cellular Cu concentrations, whereby proliferative immortality, angiogenesis and metastasis are cancer hallmarks with defined requirements for Cu. The aim of this study is to gather all known Cu-binding proteins and reveal their putative involvement in cancers using the available database resources of RNA transcript levels. Using the database along with manual curation, we identified a total of 54 Cu-binding proteins (named the human Cu proteome). Next, we retrieved RNA expression levels in cancer versus normal tissues from the TCGA database for the human Cu proteome in 18 cancer types, and noted an intricate pattern of up- and downregulation of the genes in different cancers. Hierarchical clustering in combination with bioinformatics and functional genomics analyses allowed for the prediction of cancer-related Cu-binding proteins; these were specifically inspected for the breast cancer data. Finally, for the Cu chaperone ATOX1, which is the only Cu-binding protein proposed to have transcription factor activities, we validated its predicted over-expression in patient breast cancer tissue at the protein level. This collection of Cu-binding proteins, with RNA expression patterns in different cancers, will serve as an excellent resource for mechanistic-molecular studies of Cu-dependent processes in cancer.
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| 4. |
- Bag, Pushan, 1993-
(författare)
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How could Christmas trees remain evergreen? : photosynthetic acclimation of Scots pine and Norway spruce needles during winter
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Plants and other green organisms harvest sunlight by green chlorophyll pigments and covertit to chemical energy (sugars) and oxygen in a process called photosynthesis providing the foundation for life on Earth. Although it is unanimously believed that oceanic phytoplanktons are the main contributors to the global photosynthesis, the contribution of coniferous boreal forests distributed across vast regions of the northern hemisphere cannot be undermined. Hence boreal forests account signifificantly for social, economical and environmental sustainability. Not only do conifers thrive in the tundra regions with extreme climate, but they also maintain their needles green over the boreal winter. A question remains; what makes them so resilient? In this respect, we aimed to understand the remarkable winter adaptation strategies in two dominant boreal coniferous species,i.e., Pinus sylvestris and Picea abies. First, we mapped the transcriptional landscape in Norway spruce (Picea abies) needles over the annual cycle. Transcriptional changes in the nascent needles reflflected a sequence of developmental processes and active vegetative growth during early summer and summer. Later after maturation, transcriptome reflflected activated defense against biotic factors and acclimationin response to abiotic environmental cues such as freezing temperatures during winter. Secondly, by monitoring the photosynthetic performance of Scot pine needles, we found that the trees face extreme stress during the early spring (Feb-Mar) when sub-zero temperatures are accompanied by high solar radiation. At this time, drastic changes occur in the thylakoid membranes of the chloroplast that allows the mixing of photosystem I and photosystem II that typically remain laterally segregated. This triggers direct energy transfer from PSII to PSI and thus protects PSII from damage. Furthermore, we found that this loss of lateral segregation may be a consequence of triple phosphorylationof Lhcb1 (Light harvesting complex1 of photosystem II). The structural changes in thylakoid membranes also lead to changes inthe thylakoid macro domain organisationand pigment protein composition. Furthermore, we discovered that while PSII is protected by direct energy transfer, the protection of PSI is provided through photoreduction of oxygen by flavodiiron proteins, which in turn allows P700 to stay in an oxidised state necessary for direct energy transfer. These coordinated cascades of changes concomitantly protect both PSI and PSII to maintain the needles green over the winter.
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| 5. |
- Mamontov, Eugen, 1955
(författare)
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Homeorhesis and evolutionary properties of living systems: From ordinary differential equations to the active-particle generalized kinetics theory
- 2006
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Ingår i: 10th Evolutionary Biology Meeting at Marseilles, 20-22 September 2006, Marseilles, France.
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Konferensbidrag (refereegranskat)abstract
- Advanced generalized-kinetic-theory (GKT) models for biological systems are developed for populations of active (or living) particles [1]-[5]. These particles are described with both the stochastic variables common in kinetic theory (such as time, the particle random location and velocity) and the stochastic variables related to the internal states of an active particle. Evolution of these states represents biological, ecological, or social properties of the particle behavior. Paper [6] analyzes a number of the well-known statistical-mechanics approaches and shows that the active-particle GKT (APGKT) is the only treatment capable of modelling living systems. Work [2] summarizes the significance of the notion of an active particle in kinetic models. This notion draws attention to the features distinguishing living matter from nonliving matter. They are discussed by many authors (e.g., [7]-[15], [1]-[3], [6], [16]-[18]). Work [11] considers a lot of differences between living and nonliving matters, and the limitations of the modelling approaches developed for nonliving matter. Work [6] mainly focuses on the comparison of a few theoretical mechanics treatments in terms of the key living-matter properties formulated in [15]. One of the necessary properties of the evolution of living systems is homeorhesis. It is, loosely speaking, a peculiar qualitative and quantitative insensitivity of a living system to the exogenous signals acting on it. The earlier notion, homeostasis, was introduced by W. B. Cannon in 1926 who discussed the phenomenon in detail later [7]. Homeorhesis introduced by C. H. Waddington [8, p. 32] generalizes homeostasis and is well known in biology [8], [9], [12]. It is an inherent part of mathematical models for oncogeny (e.g., [16]-[18], [6, Appendix]). Homeorhesis is also discussed in [3, Section 4] in connection with APGKT. Homeorhesis is documented in ecology (e.g., [11], [13, the left column on p. 675]) where it is one of the key notions of the strong Gaia theory, a version of the Gaia theory (e.g., [14, Chapter 8]). The strong Gaia theory “states that the planet with its life, a single living system, is regulated in certain aspects by that life” [14, p. 124]. The very origin of the name “Gaia” is related to homeorhesis or homeostasis [14, p. 118]. These notions are also used in psychology and sociology. If evolution of a system is not homeorhetic, the system can not be living. Work [6, Appendix] derives a preliminary mathematical formulation of homeorhesis in terms of the simplest dynamical systems, i.e. ordinary differential equations (ODEs). The present work complements, extended, and further specify the approach of [6, Appendix]. The work comprises the two main parts. The first part develops the sufficient conditions for ODE systems to describe homeorhesis, and suggests a fairly general structure of the ODE model. It regards homeorhesis as piecewise homeostasis. The model can be specified in different ways depending on specific systems and specific purposes of the analysis. An example of the specification is also noted (the PhasTraM nonlinear reaction-diffusion model for hyperplastic oncogeny [16]-[18]). The second part of the work discusses implementation of the above homeorhesis ODE model in terms of a special version [3] of APGKT (see above). The key feature of this version is that the components of a living population need not be discrete: the subdivision into the components is described with a general, continuous-discrete probability distribution (see also [6]). This enables certain properties of living matter noted in [15]. Moreover, the corresponding APGKT model presents a system of, firstly, a generalized kinetic equation for the conditional distribution function conditioned by the internal states of the population and, secondly, Ito's stochastic differential equations for these states. This treatement employs the results on nonstationary invariant diffusion stochastic processes [19]. The second part of the work also stresses that APGKT is substantially more important for the living-matter analysis than in the case of nonliving matter. One of the reasons is certain limitations in experimental sampling of the living-system modes presented with stochastic processes. A few directions for future research are suggested as well. REFERENCES: [1] Bellomo, N., Bellouquid, A. and Delitala, M., 2004, Mathematical topics on the modelling complex multicellular systems and tumor immune cells competition, Math. Models Methods Appl. Sci., 14, 1683-1733. [2] Bellomo, N., 2006, New hot Paper Comments, Essential Science Indicators, http://www.esi-topics.com/nhp/2006 /may- 06-NicolaBellomo.html. [3] Willander, M., Mamontov, E. and Chiragwandi, Z., 2004, Modelling living fluids with the subdivision into the components in terms of probability distributions, Math. Models Methods Appl. Sci. 14, 1495-1520. [4] Bellomo, N. and Maini, P.K., 2005, Preface and the Special Issue “Multiscale Cancer Modelling-A New Frontier in Applied Mathematics”, Math. Models Methods Appl. Sci., 15, iii-viii. [5] De Angelis, E. and Delitala, M., 2006, Modelling complex systems in applied sciences: Methods and tools of the mathematical kinetic theory for active particles. Mathl Comput. Modelling, 43, 1310-1328. [6] Mamontov, E., Psiuk-Maksymowicz, K. and Koptioug, A., 2006, Stochastic mechanics in the context of the properties of living systems, Mathl Comput. Modelling, Article in Press, 13 pp. [7] Cannon, W.B., 1932, The Wisdom of the Body (New York: Norton). [8] Waddington, C.H., 1957, The Strategy of the Genes. A Discussion of Some Aspects of Theoretical Biology (London, George Allen and Unwin). [9] Waddington, C.H., 1968, Towards a theoretical biology, Nature, 218, 525-527. [10] Cotnoir, P.-A., 1981, La compétence environnementale: Une affaire d’adaptation. Séminaire en écologie behaviorale, Univeristé du Québec, Montralé. Available online at: http://pac.cam.org/culture.doc . [11] O’Neill, R.V., DeAngelis, D.L., Waide, J.B. and Allen, T.F.H., 1986, A Hierarchical Concept of Ecosystems, Princeton: Princeton Univ. Press). [12] Sauvant, D., 1992, La modélisation systémique en nutrition, Reprod. Nutr. Dev., 32, 217-230. [13] Christensen, N.L., Bartuska, A.M., Brown, J.H., Carpenter, S., D'Antonio, C., Francis, R., Franklin, J.F., MacMahon, J.A., Noss, R.F., Parsons, D.J., Peterson, C.H., Turner, M.G. and Woodmansee, R.G., 1996, The Report of the Ecological Society of America Committee on the Scientific Basis for Ecosystem Management, Ecological Applications, 6, 665-691. Available online at: http://www.esa.org/pao/esaPositions/Papers/ReportOfSBEM.php. [14] Margulis, L., 1998, Symbiotic Planet. A New Look at Evolution (Amherst: Sciencewriters). [15] Hartwell, L.H., Hopfield, J.J., Leibler, S. and Murray, A.W., 1999, From molecular to modular cell biology, Nature, 402, C47-C52. [16] Mamontov, E., Koptioug, A.V. and Psiuk-Maksymowicz, K., 2006, The minimal, phase-transition model for the cell- number maintenance by the hyperplasia-extended homeorhesis, Acta Biotheoretica, 54, 44 pp., (no. 2, May-June, accepted). [17] Psiuk-Maksymowicz, K. and Mamontov, E., 2005, The time-slices method for rapid solving the Cauchy problem for nonlinear reaction-diffusion equations in the competition of homeorhesis with genotoxically activated hyperplasia, In: European Conference on Mathematical and Theoretical Biology - ECMTB05 (July 18-22, 2005) Book of Abstracts, Vol.1 (Dresden: Center for Information Services and High Performance Computing, Dresden Univ. Technol.), p. 429 (http://www.ecmtb05.org/). [18] Psiuk-Maksymowicz, K. and Mamontov, E., 2006, The homeorhesis-based modelling and fast numerical analysis for oncogenic hyperplasia under radiation therapy, submitted. [19] Mamontov, E., 2005, Nonstationary invariant distributions and the hydrodynamic-style generalization of the Kolmogorov-forward/Fokker-Planck equation, Appl. Math. Lett. 18 (9) 976-982.
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| 6. |
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| 7. |
- Wieloch, Thomas, 1979-
(författare)
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Intramolecular isotope analysis reveals plant ecophysiological signals covering multiple timescales
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Our societies' wellbeing relies on stable and healthy environments. However, our current lifestyles, growth-oriented economic policies and the population explosion are leading to potentially catastrophic degradation of ecosystems and progressive disruption of food chains. Hopefully, more clarity about what the future holds in store will trigger stronger efforts to find, and adopt, problem-focused coping strategies and encourage environmentally friendly lifestyles.Forecasting environmental change/destruction is complicated (inter alia) by lack of complete understanding of plant-environment interactions, particularly those involved in slow processes such as plant acclimatisation and adaptation. This stems from deficiencies in tools to analyse such slow processes. The present work aims at developing tools that can provide retrospective ecophysiological information covering timescales from days to millennia.Natural archives, such as tree-rings, preserve plant metabolites over long timescales. Analyses of intramolecular isotope abundances in plant metabolites have the potential to provide retrospective information about metabolic processes and underlying environmental controls. Thus, my colleagues and I (hereafter we) analysed intramolecular isotope patterns in tree rings to develop analytical tools that can convey information about clearly-defined plant metabolic processes over multiple timescales. Such tools might help (inter alia) to constrain plants' capacities to sequester excess amounts of anthropogenic CO2; the so-called CO2 fertilisation effect. This, in turn, might shed light on plants' sink strength for the greenhouse gas CO2, and future plant performance and growth under climate change.In the first of three studies, reported in appended papers, we analysed intramolecular 13C/12C ratios in tree-ring glucose. In six angiosperm and six gymnosperm species we found pronounced intramolecular 13C/12C differences, exceeding 10‰. These differences are transmitted into major global C pools, such as soil organic matter. Taking intramolecular 13C/12C differences into account might improve isotopic characterisation of soil metabolic processes and soil CO2 effluxes. In addition, we analysed intramolecular 13C/12C ratios in a Pinus nigra tree-ring archive spanning the period 1961 to 1995. These data revealed new ecophysiological 13C/12C signals, which can facilitate climate reconstructions and assessments of plant-environment interactions at higher resolution; thus providing higher quality information. We proposed that 13C/12C signals at glucose C-1 to C-2 derive from carbon injection into the Calvin-Benson cycle via the oxidative pentose phosphate pathway. We concluded that intramolecular 13C/12C measurements provide valuable new information about long-term metabolic dynamics for application in biogeochemistry, plant physiology, plant breeding, and paleoclimatology.In the second study, we developed a comprehensive theory on the metabolic and ecophysiological origins of 13C/12C signals at tree-ring glucose C-5 and C-6. According to this theory and theoretical implications of the first study on signals at C-1 to C-3, analysis of such intramolecular signals can provide information about several metabolic processes. At C-3, a well-known signal reflecting CO2 uptake is preserved. The glucose-6-phosphate shunt around the Calvin-Benson cycle affects 13C/12C compositions at C-1 and C-2, while the 13C/12C signals at C-5 and C-6 reflect carbon fluxes into downstream metabolism. This theoretical framework enables further experimental studies to be conducted in a hypothesis-driven manner. In conclusion, the intramolecular approach provides information about carbon allocation in plant leaves. Thus, it gives access to long-term information on key ecophysiological processes, which could not be acquired by previous approaches.The abundance of the hydrogen isotope deuterium, δD, is important for linking the water cycle with plant ecophysiology. The main factors affecting δD in plant organic matter are commonly assumed to be the δD in source water and leaf-level evaporative enrichment. Current δD models incorporate biochemical D fractionations as constants. In the third study we showed that biochemical D fractionations respond strongly to low ambient CO2 levels and low light intensity. Thus, models of δD values in plant organic matter should incorporate biochemical fractionations as variables. In addition, we found pronounced leaf-level δD differences between α-cellulose and wax n-alkanes. We explained this by metabolite-specific contributions of distinct hydrogen sources during biosynthesis.Overall, this work advances our understanding of isotope distributions and isotope fractionations in plants. It reveals the immense potential of intramolecular isotope analyses for retrospective assessment of plant metabolism and associated environmental controls.
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| 8. |
- Ohrvik, Helena, et al.
(författare)
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Identification of New Potential Interaction Partners for Human Cytoplasmic Copper Chaperone Atox1: Roles in Gene Regulation?
- 2015
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 16:8, s. 16728-39
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Tidskriftsartikel (refereegranskat)abstract
- The human copper (Cu) chaperone Atox1 delivers Cu to P1B type ATPases in the Golgi network, for incorporation into essential Cu-dependent enzymes. Atox1 homologs are found in most organisms; it is a 68-residue ferredoxin-fold protein that binds Cu in a conserved surface-exposed Cys-X-X-Cys (CXXC) motif. In addition to its well-documented cytoplasmic chaperone function, in 2008 Atox1 was suggested to have functionality in the nucleus. To identify new interactions partners of Atox1, we performed a yeast two-hybrid screen with a large human placenta library of cDNA fragments using Atox1 as bait. Among 98 million fragments investigated, 25 proteins were found to be confident interaction partners. Nine of these were uncharacterized proteins, and the remaining 16 proteins were analyzed by bioinformatics with respect to cell localization, tissue distribution, function, sequence motifs, three-dimensional structures and interaction networks. Several of the hits were eukaryotic-specific proteins interacting with DNA or RNA implying that Atox1 may act as a modulator of gene regulation. Notably, because many of the identified proteins contain CXXC motifs, similarly to the Cu transport reactions, interactions between these and Atox1 may be mediated by Cu.
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| 9. |
- Ariöz, Candan, 1983-
(författare)
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Exploring the Interplay of Lipids and Membrane Proteins
- 2014
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The interplay between lipids and membrane proteins is known to affect membrane protein topology and thus have significant effect (control) on their functions. In this PhD thesis, the influence of lipids on the membrane protein function was studied using three different membrane protein models.A monotopic membrane protein, monoglucosyldiacylglyecerol synthase (MGS) from Acholeplasma laidlawii is known to induce intracellular vesicles when expressed in Escherichia coli. The mechanism leading to this unusual phenomenon was investigated by various biochemical and biophysical techniques. The results indicated a doubling of lipid synthesis in the cell, which was triggered by the selective binding of MGS to anionic lipids. Multivariate data analysis revealed a good correlation with MGS production. Furthermore, preferential anionic lipid sequestering by MGS was shown to induce a different fatty acid modeling of E. coli membranes. The roles of specific lipid binding and the probable mechanism leading to intracellular vesicle formation were also investigated.As a second model, a MGS homolog from Synechocystis sp. PCC6803 was selected. MgdA is an integral membrane protein with multiple transmembrane helices and a unique membrane topology. The influence of different type of lipids on MgdA activity was tested with different membrane fractions of Synechocystis. Results indicated a very distinct profile compared to Acholeplasma laidlawii MGS. SQDG, an anionic lipid was found to be the species of the membrane that increased the MgdA activity 7-fold whereas two other lipids (PG and PE) had only minor effects on MgdA. Additionally, a working model of MgdA for the biosynthesis and flow of sugar lipids between Synechocystis membranes was proposed.The last model system was another integral membrane protein with a distinct structure but also a different function. The envelope stress sensor, CpxA and its interaction with E. coli membranes were studied. CpxA autophosphorylation activity was found to be positively regulated by phosphatidylethanolamine and negatively by anionic lipids. In contrast, phosphorylation of CpxR by CpxA revealed to be increased with PG but inhibited by CL. Non-bilayer lipids had a negative impact on CpxA phosphotransfer activity.Taken together, these studies provide a better understanding of the significance of the interplay of lipids and model membrane proteins discussed here.
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| 10. |
- Larsson, Daniel, 1981-
(författare)
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Exploring the Molecular Dynamics of Proteins and Viruses
- 2012
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Knowledge about structure and dynamics of the important biological macromolecules — proteins, nucleic acids, lipids and sugars — helps to understand their function. Atomic-resolution structures of macromolecules are routinely captured with X-ray crystallography and other techniques. In this thesis, simulations are used to explore the dynamics of the molecules beyond the static structures.Viruses are machines constructed from macromolecules. Crystal structures of them reveal little to no information about their genomes. In simulations of empty capsids, we observed a correlation between the spatial distribution of chloride ions in the solution and the position of RNA in crystals of satellite tobacco necrosis virus (STNV) and satellite tobacco mosaic virus (STMV). In this manner, structural features of the non-symmetric RNA could also be inferred.The capsid of STNV binds calcium ions on the icosahedral symmetry axes. The release of these ions controls the activation of the virus particle upon infection. Our simulations reproduced the swelling of the capsid upon removal of the ions and we quantified the water permeability of the capsid. The structure and dynamics of the expanded capsid suggest that the disassembly is initiated at the 3-fold symmetry axis.Several experimental methods require biomolecular samples to be injected into vacuum, such as mass-spectrometry and diffractive imaging of single particles. It is therefore important to understand how proteins and molecule-complexes respond to being aerosolized. In simulations we mimicked the dehydration process upon going from solution into the gas phase. We find that two important factors for structural stability of proteins are the temperature and the level of residual hydration. The simulations support experimental claims that membrane proteins can be protected by a lipid micelle and that a non-membrane protein could be stabilized in a reverse micelle in the gas phase. A water-layer around virus particles would impede the signal in diffractive experiments, but our calculations estimate that it should be possible to determine the orientation of the particle in individual images, which is a prerequisite for three-dimensional reconstruction.
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| 11. |
- Lindholm, Ljubica, et al.
(författare)
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Effect of lipid bilayer properties on the photocycle of green proteorhodopsin
- 2015
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Ingår i: Biochimica et Biophysica Acta - Bioenergetics. - : Elsevier BV. - 0005-2728 .- 1879-2650. ; 1847:8, s. 698-708
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Tidskriftsartikel (refereegranskat)abstract
- The significance of specific lipids for proton pumping by the bacterial rhodopsin proteorhodopsin (pR) was studied. To this end, it was examined whether pR preferentially binds certain lipids and whether molecular properties of the lipid environment affect the photocycle. pR's photocyde was followed by microsecond flash-photolysis in the visible spectral range. It was fastest in phosphatidylcholine liposomes (soy bean lipid), intermediate in 3-[(3-cholamidopropyl) dimethylammonio] propanesulfonate (CHAPS): 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) bicelles and in Triton X-100, and slowest when pR was solubilized in CHAPS. In bicelles with different lipid compositions, the nature of the head groups, the unsaturation level and the fatty acid chain length had small effects on the photocycle. The specific affinity of pR for lipids of the expression host Eschetichia coil was investigated by an optimized method of lipid isolation from purified membrane protein using two different concentrations of the detergent N-dodecyl-beta-D-maltoside (DDM). We found that 11 lipids were copurified per pR molecule at 0.1% DDM, whereas essentially all lipids were stripped off from pR by 1% DDM. The relative amounts of copurifled phosphatidylethanolamine, phosphatidylglycerol, and cardiolipin did not correlate with the molar percentages normally present in E. coil cells. The results indicate a predominance of phosphatidylethanolamine species in the lipid annulus around recombinant pR that are less polar than the dominant species in the cell membrane of the expression host E. coli.
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| 12. |
- Matson Dzebo, Maria, 1985, et al.
(författare)
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Extended functional repertoire for human copper chaperones
- 2016
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Ingår i: Biomolecular Concepts. - : Walter de Gruyter GmbH. - 1868-5021 .- 1868-503X. ; 7:1, s. 29-39
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Tidskriftsartikel (refereegranskat)abstract
- Copper (Cu) ions are cofactors in many essential enzymes. As free Cu ions are toxic, most organisms have highly specialized Cu transport systems involving dedicated proteins. The human cytoplasmic Cu chaperone Atox1 delivers Cu to P1B-type ATPases in the Golgi network, for incorporation into Cu-dependent enzymes following the secretory path. Atox1 homologs are found in most organisms; it is a 68-residue ferredoxin-fold protein that binds Cu in a conserved surface-exposed CXXC motif. In addition to Atox1, the human cytoplasm also contains Cu chaperones for loading of superoxide dismutase 1 (i.e. CCS) and cytochrome c oxidase in mitochondria (i.e. Cox17). Many mechanistic aspects have been resolved with respect to how Cu ions are moved between these proteins. In addition to the primary cytoplasmic Cu chaperone function, all three cytoplasmic chaperones have been reported to have other interaction partners that are involved in signaling pathways that modulate cell growth and development. These new discoveries imply that humans have evolved a highly sophisticated network of control mechanisms that connect Cu transport with cell regulatory processes. This knowledge may eventually be exploited for future drug developments towards diseases such as cancer and neurodegenerative disorders.
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| 13. |
- Sahin, Cagla, et al.
(författare)
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Structural Basis for Dityrosine-Mediated Inhibition of α-Synuclein Fibrillization
- 2022
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 144:27, s. 11949-11954
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Tidskriftsartikel (refereegranskat)abstract
- α-Synuclein (α-Syn) is an intrinsically disordered protein which self-assembles into highly organized β-sheet structures that accumulate in plaques in brains of Parkinson’s disease patients. Oxidative stress influences α-Syn structure and self-assembly; however, the basis for this remains unclear. Here we characterize the chemical and physical effects of mild oxidation on monomeric α-Syn and its aggregation. Using a combination of biophysical methods, small-angle X-ray scattering, and native ion mobility mass spectrometry, we find that oxidation leads to formation of intramolecular dityrosine cross-linkages and a compaction of the α-Syn monomer by a factor of √2. Oxidation-induced compaction is shown to inhibit ordered self-assembly and amyloid formation by steric hindrance, suggesting an important role of mild oxidation in preventing amyloid formation.
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| 14. |
- Sridhara, Sagar, 1989
(författare)
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Multiple structural flavors of RNase P in precursor tRNA processing.
- 2024
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Ingår i: Wiley interdisciplinary reviews. RNA. - 1757-7012. ; 15:2
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Tidskriftsartikel (refereegranskat)abstract
- The precursor transfer RNAs (pre-tRNAs) require extensive processing to generate mature tRNAs possessing proper fold, structural stability, and functionality required to sustain cellular viability. The road to tRNA maturation follows an ordered process: 5'-processing, 3'-processing, modifications at specific sites, if any, and 3'-CCA addition before aminoacylation and recruitment to the cellular protein synthesis machinery. Ribonuclease P (RNase P) is a universally conserved endonuclease in all domains of life, performing the hydrolysis of pre-tRNA sequences at the 5' end by the removal of phosphodiester linkages between nucleotides at position -1 and +1. Except for an archaeal species: Nanoarchaeum equitans where tRNAs are transcribed from leaderless-position +1, RNase P is indispensable for life and displays fundamental variations in terms of enzyme subunit composition, mechanism of substrate recognition and active site architecture, utilizing in all cases a two metal ion-mediated conserved catalytic reaction. While the canonical RNA-based ribonucleoprotein RNase P has been well-known to occur in bacteria, archaea, and eukaryotes, the occurrence of RNA-free protein-only RNase P in eukaryotes and RNA-free homologs of Aquifex RNase P in prokaryotes has been discovered more recently. This review aims to provide a comprehensive overview of structural diversity displayed by various RNA-based and RNA-free RNase P holoenzymes towards harnessing critical RNA-protein and protein-protein interactions in achieving conserved pre-tRNA processing functionality. Furthermore, alternate roles and functional interchangeability of RNase P are discussed in the context of its employability in several clinical and biotechnological applications. This article is categorized under: RNA Processing > tRNA Processing RNA Evolution and Genomics > RNA and Ribonucleoprotein Evolution RNA Interactions with Proteins and Other Molecules > RNA-Protein Complexes.
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| 15. |
- Vajda, Vivi, et al.
(författare)
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Geochemical Fingerprints of Ginkgoales Across the Triassic-Jurassic Boundary of Greenland
- 2021
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Ingår i: International journal of plant sciences. - Chicago : University of Chicago Press. - 1058-5893 .- 1537-5315. ; 182:7, s. 649-662
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Tidskriftsartikel (refereegranskat)abstract
- Premise of research. Geochemical fingerprinting of fossil plants is a relatively new research field complementing morphological analyses and providing information for paleoenvironmental interpretations. Ginkgoales contains a single extant species but was diverse through the Mesozoic and is an excellent target for biochemical analyses.Methodology. Cuticles derived from fresh and fallen autumn leaves of extant Ginkgo biloba and seven fossil gink- goalean leaf taxa, one seed fern taxon, and two taxa with bennettitalean affinity were analyzed by infrared (IR) microspec- troscopy at the D7 beamline in the MAX IV synchrotron laboratory, Sweden. The fossil material derives from Triassic and Jurassic successions of Greenland. Spectral data sets were compared and evaluated by hierarchical cluster analysis (HCA) and principal component analysis performed on vector-normalized, first-derivative IR absorption spectra.Pivotal results. The IR absorption spectra of the fossil leaves all reveal signatures that clearly indicate the pres- ence of organic compounds. Spectra of the extant G. biloba leaves reveal the presence of aliphatic chains, aromatic and ester carbonyl functional groups from polymer cutin and other waxy compounds, and polysaccharides. Inter- estingly, both the extant autumn leaves and the fossil specimens reveal the presence of carboxyl/ketone molecules, suggesting that chemical alterations during the initial stages of decomposition are preserved through fossilization. Two major subclusters were identified through HCA of the fossil spectra.Conclusions. Consistent chemical IR signatures, specific for each fossil taxon are present in cuticles, and suf- ficient molecular content is preserved in key regions to reflect the plants’ original chemical signatures. The alter- ations of the organic compounds are initiated as soon as the leaves are shed, with loss of proteins and increased ester and carboxyl/ketone compound production in the fallen leaves. We further show that the groupings of taxa reflect a combination of phylogeny and environmental conditions related to the end-Triassic event.
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| 16. |
- Wu, Fei, 1993-
(författare)
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Exploring Membrane Proteins within the Inner Mitochondrial and Endoplasmic Reticulum Membranes : Mitochondrial respiratory complexes and ER-localized Shr3
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Membrane proteins play important roles in various life processes, for example, those in the inner mitochondrial membrane (IMM), endoplasmic reticulum (ER) membrane, and plasma membrane (PM). Oxidative phosphorylation complexes, densely packed in the IMM are crucial for energy transduction in eukaryotes. We determined three entire II2III2 IV2 supercomplex (SC) structures with 114 lipids at 2.1-2.4 Å resolution in Perkinsus marinus (P. marinus). The structures show a complete electron transfer pathway from complex II (CII) to complex IV (CIV). These architectures also reveal rotation states of the iron sulfur protein (ISP) in complex III (CIII), from one of which we observed two novel proteins that might impair the electron transfer. We also studied how the salt concentration and detergent affect the electron transfer. We determined the SC III2 IV-cytochrome c (cyt. c) cryo-EM structure at 20 mM salt concentration condition. Together with kinetic study, these data implicate that multiple cyt. c are involved in electron transfer between CIII and CIV. Our kinetic studies of CIV also indicate a native ligand bound near its K proton pathway which could be removed by detergent, leading to an increase in electron transfer rate and the activity of the enzyme. Most biogenesis of integral membrane proteins in eukaryotes is done in ER, such as the amino acid permeases (AAP), which function as amino acid transporters in the PM. Its synthesis and functional folding in Saccharomyces cerevisiae (S. cerevisiae) requires an ER membrane-localized chaperone, Shr3. We utilized a yeast growth-based genetic assay, in conjunction with a split-ubiquitin yeast two-hybrid assay, to demonstrate the selective interaction between Shr3 and nested C-terminal AAP truncations. This interaction exhibited a distinct pattern, wherein it gradually intensified and then weakened as more transmembrane helices folded. The work presented in this thesis contributions to an increased understanding of the organization and function of SCs, the effects of protein subunits, salt concentrations, and detergents on electron transfer, as well as the mechanism of Shr3 on AAP folding in the ER membrane. Together, these works have shed light on the understanding of the structure and function of several membrane proteins.
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| 17. |
- Devkota, Ranjan, et al.
(författare)
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The adiponectin receptor AdipoR2 and its Caenorhabditis elegans homolog PAQR-2 prevent membrane rigidification by exogenous saturated fatty acids.
- 2017
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Ingår i: PLoS genetics. - : Public Library of Science (PLoS). - 1553-7404. ; 13:9
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Tidskriftsartikel (refereegranskat)abstract
- Dietary fatty acids can be incorporated directly into phospholipids. This poses a specific challenge to cellular membranes since their composition, hence properties, could greatly vary with different diets. That vast variations in diets are tolerated therefore implies the existence of regulatory mechanisms that monitor and regulate membrane compositions. Here we show that the adiponectin receptor AdipoR2, and its C. elegans homolog PAQR-2, are essential to counter the membrane rigidifying effects of exogenously provided saturated fatty acids. In particular, we use dietary supplements or mutated E. coli as food, together with direct measurements of membrane fluidity and composition, to show that diets containing a high ratio of saturated to monounsaturated fatty acids cause membrane rigidity and lethality in the paqr-2 mutant. We also show that mammalian cells in which AdipoR2 has been knocked-down by siRNA are unable to prevent the membrane-rigidifying effects of palmitic acid. We conclude that the PAQR-2 and AdipoR2 proteins share an evolutionarily conserved function that maintains membrane fluidity in the presence of exogenous saturated fatty acids.
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| 18. |
- Hemsworth, Glyn R., et al.
(författare)
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Structural dissection of a complex Bacteroides ovatus gene locus conferring xyloglucan metabolism in the human gut
- 2016
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Ingår i: Open Biology. - : Royal Society of London. - 2046-2441. ; 6:7
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Tidskriftsartikel (refereegranskat)abstract
- The human gastrointestinal tract harbours myriad bacterial species, collectively termed the microbiota, that strongly influence human health. Symbiotic members of our microbiota play a pivotal role in the digestion of complex carbohydrates that are otherwise recalcitrant to assimilation. Indeed, the intrinsic human polysaccharide-degrading enzyme repertoire is limited to various starch-based substrates; more complex polysaccharides demand microbial degradation. Select Bacteroidetes are responsible for the degradation of the ubiquitous vegetable xyloglucans (XyGs), through the concerted action of cohorts of enzymes and glycan-binding proteins encoded by specific xyloglucan utilization loci (XyGULs). Extending recent (meta) genomic, transcriptomic and biochemical analyses, significant questions remain regarding the structural biology of the molecular machinery required for XyG saccharification. Here, we reveal the three-dimensional structures of an alpha-xylosidase, a beta-glucosidase, and two alpha-L-arabinofuranosidases from the Bacteroides ovatus XyGUL. Aided by bespoke ligand synthesis, our analyses highlight key adaptations in these enzymes that confer individual specificity for xyloglucan side chains and dictate concerted, stepwise disassembly of xyloglucan oligosaccharides. In harness with our recent structural characterization of the vanguard endo-xyloglucanse and cell-surface glycan-binding proteins, the present analysis provides a near-complete structural view of xyloglucan recognition and catalysis by XyGUL proteins.
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| 19. |
- Lindahl, Lina, 1984
(författare)
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Towards membrane engineering as a tool in cell factory design: A case study on acetic acid tolerance in Saccharomyces cerevisiae
- 2017
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The sustainable production of fuels, chemicals, and materials using renewable resources is a necessity if we are to reduce our ecological footprint and the rate of climate change. Lignocellulosic biomass, the major constituent of plant cell walls, is a renewable raw material with great potential due to its high abundance. The conversion of lignocellulosic material into desired products using microorganisms is a promising option, although many microbial production processes fail to reach the titers required for process economy due to cellular inhibition. The inhibitory action of some compounds is related to the physiochemical properties of the cell membrane. Inhibitors may enter the cell by passive diffusion through the lipid bilayer of the cell membrane, or may inhibit cells by partition in the lipid bilayer, altering the membrane properties. The aim of the research described in this thesis was to evaluate the possibility of engineering the lipid composition of the cell membrane to create microbial cell factories with maintained production capacity when exposed to compounds whose mechanism of inhibition relates to the physiochemical properties of the membrane. Attempts were made to increase the tolerance of Saccharomyces cerevisiae to the lignocellulose-derived inhibitor acetic acid, by engineering the cell membrane in order to reduce the rate of acetic acid diffusion. Studies of the acetic-acid-tolerant yeast Zygosaccharomyces bailii revealed that its high tolerance relies on its ability to remodel the cell membrane lipid composition so as to greatly increase the fraction of sphingolipids. Further evidence that sphingolipids reduce the rate of acetic acid diffusion was obtained by molecular dynamics simulations of model membranes with increasing fraction of sphingolipids. The lipid metabolism of S. cerevisiae was then engineered in an attempt to increase the fraction of sphingolipids in the cell membrane. However, sphingolipid synthesis was unchanged or decreased in these strains. The effect of sphingolipids on acetic acid tolerance in S. cerevisiae could therefore not be elucidated, but insight was gained into sphingolipid regulation. To understand the variation in membrane permeation, in particular the extent to which compounds partitioning in the cell membrane change the rate of acetic acid diffusion, the effects of ethanol and n-butanol were investigated. It was found that target titers in ethanol and n‑butanol production significantly increased the rate of acetic acid diffusion; n-butanol having a stronger effect than ethanol. Molecular dynamics simulations were then used to suggest mechanisms for the experimental observations.
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| 20. |
- Lorentzon, Emma, 1995, et al.
(författare)
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Effects of the toxic metals arsenite and cadmium on α-synuclein aggregation in vitro and in cells
- 2021
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 22:21
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Tidskriftsartikel (refereegranskat)abstract
- Exposure to heavy metals, including arsenic and cadmium, is associated with neurodegen-erative disorders such as Parkinson’s disease. However, the mechanistic details of how these metals contribute to pathogenesis are not well understood. To search for underlying mechanisms involving α-synuclein, the protein that forms amyloids in Parkinson’s disease, we here assessed the effects of arsenic and cadmium on α-synuclein amyloid formation in vitro and in Saccharomyces cerevisiae (budding yeast) cells. Atomic force microscopy experiments with acetylated human α-synuclein demonstrated that amyloid fibers formed in the presence of the metals have a different fiber pitch compared to those formed without metals. Both metal ions become incorporated into the amyloid fibers, and cadmium also accelerated the nucleation step in the amyloid formation process, likely via binding to intermediate species. Fluorescence microscopy analyses of yeast cells expressing fluorescently tagged α-synuclein demonstrated that arsenic and cadmium affected the distribution of α-synuclein aggregates within the cells, reduced aggregate clearance, and aggravated α-synuclein toxicity. Taken together, our in vitro data demonstrate that interactions between these two metals and α-synuclein modulate the resulting amyloid fiber structures, which, in turn, might relate to the observed effects in the yeast cells. Whilst our study advances our understanding of how these metals affect α-synuclein biophysics, further in vitro characterization as well as human cell studies are desired to fully appreciate their role in the progression of Parkinson’s disease.
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| 21. |
- Skoog, Emma, 1983
(författare)
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Biobased Adipic Acid - Challenges in Establishing a Cell Factory
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Growing concern regarding climate change calls for sustainable solutions to significantly reduce our dependency on non-renewable fossil-based raw materials. One potential solution is the development of biorefineries where biobased, renewable raw materials are converted into valuable products via enzymatic, chemical or microbial conversion. This work focuses on the microbial production of adipic acid, a precursor in the nylon industry, currently derived from fossil-based raw material. No known naturally occurring microorganism is able to produce adipic acid, and genetic engineering of a suitable host is therefore required. The aim of the work presented in this thesis was to engineer a microorganism for the production of adipic acid from glucose, more specifically, from glucose streams derived from lignocellulosic forest residues. Theoretical evaluation of metabolic pathways for adipic acid production revealed several obstacles, including redox imbalance and the discovery or engineering of enzymes to catalyze novel reactions. Mining of enzyme databases for alternative paths proved fruitful, and the number of biochemical reactions in the lysine pathway employing as yet unidentified enzymes was reduced from three to two, without affecting the thermodynamics of the pathway. A combined approach of in vitro and in silico analysis suggested potential enzyme engineering strategies for one of the reactions, for which there are as yet no identified enzymes, namely, the reduction of unsaturated α,β bonds of 6-aminohex-2-enoic acid and 2-hexenedioic acid. When defining a suitable host for microbial adipic acid production, tolerance to high concentrations of adipic acid (50-100 g L-1) is important to ensure an economically feasible process, preferably at low pH (below 5) to further reduce the overall process cost. Screening of bacteria, yeasts and a filamentous fungus grown in increasing concentrations of adipic acid (0-100 g L-1) and at different pH revealed Candida viswanathii to be a promising host to engineer for adipic acid production. A comparative study of C. viswanathii with Saccharomyces cerevisiae in controlled batch cultivations at increasing adipic acid concentrations (0-95 g L-1) and low pH (pH 4 and pH 5) revealed significant differences in their tolerance to adipic acid; C. viswanathii being able to grow, almost unaffected, under all the conditions investigated, whereas S. cerevisiae was unable to grow at 95 g L-1. Lipid analysis of their cell membranes revealed C. viswanathii to have a thicker and more compact cell membrane, which is probably less permeable to adipic acid.
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| 22. |
- Wu, Min, 1986, et al.
(författare)
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Proline 411 biases the conformation of the intrinsically disordered plant UVR8 photoreceptor C27 domain altering the functional properties of the peptide
- 2019
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 9
-
Tidskriftsartikel (refereegranskat)abstract
- UVR8 (UV RESISTANCE LOCUS 8) is a UV-B photoreceptor responsible for initiating UV-B signalling in plants. UVR8 is a homodimer in its signalling inactive form. Upon absorption of UV radiation, the protein monomerizes into its photoactivated state. In the monomeric form, UVR8 binds the E3 ubiquitin ligase COP1 (CONSTITUTIVELY PHOTOMORPHOGENIC 1), triggering subsequent UV-B-dependent photomorphogenic development in plants. Recent in vivo experiments have shown that the UVR8 C-terminal region (aa 397-423; UVR8(C27)) alone is sufficient to regulate the activity of COP1. In this work, CD spectroscopy and NMR experiments showed that the UVR8(C27) domain was non-structured but gained secondary structure at higher temperatures leading to increased order. Bias-exchange metadynamics simulations were also performed to evaluate the free energy landscape of UVR8(C27). An inverted free energy landscape was revealed, with a disordered structure in the global energy minimum. Flanking the global energy minimum, more structured states were found at higher energies. Furthermore, stabilization of the low energy disordered state was attributed to a proline residue, P411, as evident from P411A mutant data. P411 is also a key residue in UVR8 binding to COP1. UVR8(C27) is therefore structurally competent to function as a molecular switch for interaction of UVR8 with different binding partners since at higher free energies different structural conformations are being induced in this peptide. P411 has a key role for this function.
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| 23. |
- Zeng, Jiao, et al.
(författare)
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High-resolution structure of a fish aquaporin reveals a novel extracellular fold
- 2022
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Ingår i: Life Science Alliance. - : Life Science Alliance, LLC. - 2575-1077. ; 5:12
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Tidskriftsartikel (refereegranskat)abstract
- Aquaporins are protein channels embedded in the lipid bilayer in cells from all organisms on earth that are crucial for water homeostasis. In fish, aquaporins are believed to be important for osmoregulation; however, the molecular mechanism behind this is poorly understood. Here, we present the first structural and functional characterization of a fish aquaporin; cpAQP1aa from the fresh water fish climbing perch (Anabas testudineus), a species that is of high osmoregulatory interest because of its ability to spend time in seawater and on land. These studies show that cpAQP1aa is a water-specific aquaporin with a unique fold on the extracellular side that results in a constriction region. Functional analysis combined with molecular dynamic simulations suggests that phosphorylation at two sites causes structural perturbations in this region that may have implications for channel gating from the extracellular side.
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| 24. |
-
Advances in Thermal Imaging
- 2021
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Ingår i: Journal of Thermal Biology. - 0306-4565.
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Annan publikation (refereegranskat)abstract
- Thermal imaging, or more correctly infrared thermography, has been widely applied to studies of animal and human biology (see Burnay et al. 1988; McCafferty 2007; Soerensen and Pedersen 2015; Fernandez-Cuevas et al., 2015; Tattersall 2016). This technique provides non-contact measurement of surface temperature, allowing real-time recording of the spatial temperature distribution of a body region, physical structure or habitat of interest. Thermal imaging technology has advanced rapidly in the last decade and is now becoming a key tool in thermal biology. Technological advances include greater spatial and temporal resolution, increased capacity to record and store high resolution radiometric video, as well as reduced device size and portability. In addition, high-quality thermal imaging devices are quickly becoming more affordable, meaning thermal imaging is an increasingly common item of the research tool-kit in many pure and applied fields. The aim of this Special Issue was to highlight how advances in thermal imaging can be used to answer important questions in biology, and to demonstrate how the combination of this technology with novel analytical methods can further advance our conceptual understanding of thermal biology.
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| 25. |
- Ahmadpour, Doryaneh, 1973, et al.
(författare)
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Inhibition of MAPK Hog1 Results in Increased Hsp104 Aggregate Formation Probably through Elevated Arsenite Influx into the Cells, an Approach with Numerous Potential Applications
- 2014
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Ingår i: American Journal of Molecular Biology. - : Scientific Research Publishing, Inc.. - 2161-6620 .- 2161-6663. ; 4:2, s. 59-71
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Tidskriftsartikel (refereegranskat)abstract
- Arsenic is a highly toxic and carcinogenic metalloid widely dispersed in the environment, contaminating water and soil and accumulating in crops. Paradoxically, arsenic is also part of modern therapy and employed in treating numerous ailments and diseases. Hence, inventing strategies to tune cellular arsenic uptake based on purpose is striking. Here, we describe an approach in which the arsenite uptake can be increased using a MAPK inhibitor. Employing microfluidic flow chambers in combination with optical tweezers and fluorescent microscopy, we elevated the influx of arsenite into the yeast Saccharomyces cerevisiae cells following short-term treatment with a Hog1 kinase inhibitor. The increase in arsenite uptake was followed on arsenite triggered redistribution of a reporter protein, Hsp104-GFP, which was imaged over time. The effect was even more pronounced when the yeast mother and daughter cells were analyzed disjointedly, an opportunity provided owing to single-cell analysis. Our data firstly provide a strategy to increase arsenite uptake and secondly show that arsenite triggered aggregates, previously shown to be sites of damaged proteins, are distributed asymmetrically and less accumulated in daughter cells. Inventing approaches to tune arsenite uptake has a great value for its use in environmental as well as medical applications.
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