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| 2. |
- Abbas, Abdul-Karim, 1959, et al.
(författare)
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Persistent LTP without triggered protein synthesis.
- 2009
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Ingår i: Neuroscience research. - : Elsevier BV. - 0168-0102. ; 63:1, s. 59-65
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Tidskriftsartikel (refereegranskat)abstract
- Protein synthesis is believed to be involved in stabilizing synaptic plasticity. Effects lasting longer than about 2-3h are considered to require synthesis of new proteins, implying a functional separation between early (E) and late (L) components. However, the issue of constitutive vs. new protein synthesis is still unclear, especially in young animals. Here, we examined the effects of two protein synthesis inhibitors, anisomycin and emetine, on long-term-potentiation (LTP) in CA1 area of hippocampal slices from 12- to 20-day-old rats. Either drug was applied from -30 min to +30 min with respect to LTP induction, a time window previously reported to be critical. However, the LTP remained stable under the entire recording period of 4h (anisomycin), or 8h (emetine). Proper preparation of emetine solution was evidenced by the fact that, in separate experiments, prolonged treatment with emetine gradually blocked baseline responses. Although no corresponding effect was observed with anisomycin, the drug was judged to be potent by its ability to inhibit yeast growth. The ability of anisomycin to inhibit protein synthesis was further confirmed by radiolabeling experiments assessing the degree of leucine incorporation. Our data suggest that LTP up to at least 8h is not dependent on triggered protein synthesis but can be attained by utilizing proteins already available at induction time.
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| 3. |
- Alexandersson, Erik, et al.
(författare)
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Transcriptional regulation of aquaporins in accessions of Arabidopsis in response to drought stress.
- 2010
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Ingår i: Plant Journal. - 1365-313X. ; 61, s. 650-660
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Tidskriftsartikel (refereegranskat)abstract
- Summary Aquaporins facilitate water transport over cellular membranes and are therefore believed to play an important role in water homeostasis. In higher plants aquaporin-like proteins, also called major intrinsic proteins (MIPs), are divided into 5 subfamilies. We have previously shown that MIP transcription in Arabidopsis thaliana generally is down-regulated in leaves upon drought stress, apart from two members of the Plasma membrane Intrinsic Protein (PIP) subfamily, AtPIP1;4 and AtPIP2;5, which are up-regulated. In order to assess if this regulation is general or accession-specific we monitored gene expression of all PIPs in five Arabidopsis accessions. Overall drought regulation of PIPs was well conserved for all five accessions tested suggesting a general and fundamental physiological role of this drought response. In addition, significant differences among accessions were identified for transcripts of three PIP genes. Principal component analysis showed that most of the PIP transcriptional variation during drought stress could be explained by one variable linked to leaf water content. Promoter-GUS constructs of AtPIP1;4, AtPIP2;5 and also AtPIP2;6, which is unresponsive to drought stress, had distinct expression patterns concentrated to the base of the leaf petioles and parts of the flowers. The presence of drought stress response elements within the 1.6 kb promoter regions of AtPIP1;4 and AtPIP2;5, was demonstrated by comparing transcription of the promoter reporter construct and the endogenous gene upon drought stress. Analysis by ATTED-II and other web-based bioinformatical tools showed that several of the MIPs down-regulated upon drought are strongly co-expressed, whereas AtPIP1;4, AtPIP2;5 and AtPIP2;6 are not co-expressed.
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| 4. |
- Anderberg, Hanna, et al.
(författare)
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Algal MIPs, high diversity and conserved motifs
- 2011
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Ingår i: BMC Evolutionary Biology. - : Springer Science and Business Media LLC. - 1471-2148. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Background: Major intrinsic proteins (MIPs) also named aquaporins form channels facilitating the passive transport of water and other small polar molecules across membranes. MIPs are particularly abundant and diverse in terrestrial plants but little is known about their evolutionary history. In an attempt to investigate the origin of the plant MIP subfamilies, genomes of chlorophyte algae, the sister group of charophyte algae and land plants, were searched for MIP encoding genes. Results: A total of 22 MIPs were identified in the nine analysed genomes and phylogenetic analyses classified them into seven subfamilies. Two of these, Plasma membrane Intrinsic Proteins (PIPs) and GlpF-like Intrinsic Proteins (GIPs), are also present in land plants and divergence dating support a common origin of these algal and land plant MIPs, predating the evolution of terrestrial plants. The subfamilies unique to algae were named MIPA to MIPE to facilitate the use of a common nomenclature for plant MIPs reflecting phylogenetically stable groups. All of the investigated genomes contained at least one MIP gene but only a few species encoded MIPs belonging to more than one subfamily. Conclusions: Our results suggest that at least two of the seven subfamilies found in land plants were present already in an algal ancestor. The total variation of MIPs and the number of different subfamilies in chlorophyte algae is likely to be even higher than that found in land plants. Our analyses indicate that genetic exchanges between several of the algal subfamilies have occurred. The PIP1 and PIP2 groups and the Ca2+ gating appear to be specific to land plants whereas the pH gating is a more ancient characteristic shared by all PIPs. Further studies are needed to discern the function of the algal specific subfamilies MIPA-E and to fully understand the evolutionary relationship of algal and terrestrial plant MIPs.
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| 5. |
- Danielson, Jonas, et al.
(författare)
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Phylogeny of Major Intrinsic Proteins
- 2010
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Ingår i: Advances in Experimental Medicine and Biology. - 0065-2598. ; 679, s. 19-32
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Tidskriftsartikel (refereegranskat)abstract
- Major intrinsic proteins (MIPs) form a large superfamily of proteins that can be divided into different subfamilies and groups according to phylogenetic analyses. Plants encode more MIPs than other organisms and seven subfamilies have been defined, whereof the Nodulin26-like major intrinsic proteins (NIPs) have been shown to permeate metalloids. In this chapter we review the phylogeny of MIPs in general and especially of the plant MIPs. We also identify bacterial NIP-like MIPs and discuss the evolutionary implications of this finding regarding the origin and ancestral transport specificity of the NIPs.
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| 6. |
- Danielson, Jonas
(författare)
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Plant Major Intrinsic Proteins - natural variation and evolution
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Major Intrinsic Proteins (MIPs, also called Aquaporins, AQPs) are channel forming membrane proteins. Although initially functionally characterized and named after their water channeling property in human red blood cells, it has become increasingly evident that MIPs are present in all types of organisms and transport a variety of small, uncharged molecules besides water. MIPs have a highly conserved structure with a constriction region and electrostatic repulsion filter allowing the combination of high transport rate and selectivity, characteristic of MIPs. In plants, MIPs form a large and varied protein family, with roughly three times as many isoforms as found in animals. Even though the abundance implies that MIPs have important functions in plants, the roles of individual MIPs have so far only been described for a handful of isoforms. Variations in the filter regions as well as experimental data, suggest differences in substrate specificities and localization for different MIP subfamilies. However actual functions of different isoforms remain largely unknown as traditional knock out/knock down experiments to a large extent have failed to reveal any clear phenotypes. Using another approach, we used naturally occurring genetic variants (accessions) of the model plant Arabidopsis thaliana to see if differences in traits, such as drought tolerance, could be linked to differences in MIP regulation. We also looked at the evolution of the MIP family in plants, to see if this could for example be linked to events such as the emergence of a vascular system or the development of a terrestrial lifestyle. Therefore we identified the whole MIP family in the moss Physcomitrella patens and in nine different green algae and investigated how these relate to those of higher plants. An amazing diversity of MIPs was found and surprisingly some of the higher plant subfamilies were present also in these simple plants.
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| 7. |
- Danielson, Jonas, et al.
(författare)
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Unexpected complexity of the Aquaporin gene family in the moss Physcomitrella patens
- 2008
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Ingår i: BMC Plant Biology. - : Springer Science and Business Media LLC. - 1471-2229. ; 8:45
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Tidskriftsartikel (refereegranskat)abstract
- Background: Aquaporins, also called major intrinsic proteins (MIPs), constitute an ancient superfamily of channel proteins that facilitate the transport of water and small solutes across cell membranes. MIPs are found in almost all living organisms and are particularly abundant in plants where they form a divergent group of proteins able to transport a wide selection of substrates. Results: Analyses of the whole genome of Physcomitrella patens resulted in the identification of 23 MIPs, belonging to seven different subfamilies, of which only five have been previously described. Of the newly discovered subfamilies one was only identified in P. patens (Hybrid Intrinsic Protein, HIP) whereas the other was found to be present in a wide variety of dicotyledonous plants and forms a major previously unrecognized MIP subfamily (X Intrinsic Proteins, XIPs). Surprisingly also some specific groups within subfamilies present in Arabidopsis thaliana and Zea mays could be identified in P. patens. Conclusion: Our results suggest an early diversification of MIPs resulting in a large number of subfamilies already in primitive terrestrial plants. During the evolution of higher plants some of these subfamilies were subsequently lost while the remaining subfamilies expanded and in some cases diversified, resulting in the formation of more specialized groups within these subfamilies.
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| 8. |
- Jood, Katarina, 1966, et al.
(författare)
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Fibrinogen gene variation and ischemic stroke.
- 2008
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Ingår i: Journal of thrombosis and haemostasis : JTH. - : Elsevier BV. - 1538-7836. ; 6:6, s. 897-904
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Plasma fibrinogen level and fibrin clot structure are heritable traits that may be of importance in the pathogenesis of ischemic stroke. OBJECTIVES: To investigate associations between variation in the fibrinogen gamma (FGG), alpha (FGA) and beta (FGB) genes, fibrinogen level, and ischemic stroke. METHODS: The Sahlgrenska Academy Study on Ischemic Stroke comprises 600 cases and 600 matched population controls. Stroke subtypes were defined according to TOAST criteria. Plasma fibrinogen level was measured by an automated clot-rate assay. Eight tagging single nucleotide polymorphisms (SNPs) were selected to capture genetic variation in the FGA, FGG, and FGB genes. RESULTS: Plasma fibrinogen was independently associated with overall ischemic stroke and all subtypes, both in the acute stage (P < 0.001) and at three-month follow-up (P < 0.05). SNPs belonged to two haplotype blocks, one containing the FGB gene and the other the FGG and FGA genes. FGB haplotypes were associated with fibrinogen level (P < 0.01), but not with ischemic stroke. In contrast, FGG/FGA haplotypes showed independent association to ischemic stroke but not to fibrinogen level. In an additive model with the most common FGG/FGA haplotype (A1) as reference, the adjusted odds ratios of ischemic stroke were 1.4 [95% confidence interval (95% CI) 1.1-1.8], P < 0.01, 1.4 (95% CI 1.0-1.8), P < 0.05, and 1.5 (95% CI 1.0-2.1), P < 0.05 for the A2, A3, and A4 FGG/FGA haplotypes, respectively. CONCLUSION: FGG/FGA haplotypes show association to ischemic stroke. This association is independent of fibrinogen level, thus suggesting that the association between ischemic stroke and variation at the FGG/FGA genes is mediated by qualitative rather than quantitative effects on fibrin(ogen).
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| 9. |
- Nordén, Kristina, et al.
(författare)
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Increasing gene dosage greatly enhances recombinant expression of aquaporins in Pichia pastoris
- 2011
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Ingår i: BMC Biotechnology. - : Springer Science and Business Media LLC. - 1472-6750. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Background: When performing functional and structural studies, large quantities of pure protein are desired. Most membrane proteins are however not abundantly expressed in their native tissues, which in general rules out purification from natural sources. Heterologous expression, especially of eukaryotic membrane proteins, has also proven to be challenging. The development of expression systems in insect cells and yeasts has resulted in an increase in successful overexpression of eukaryotic proteins. High yields of membrane protein from such hosts are however not guaranteed and several, to a large extent unexplored, factors may influence recombinant expression levels. In this report we have used four isoforms of aquaporins to systematically investigate parameters that may affect protein yield when overexpressing membrane proteins in the yeast Pichia pastoris. Results: By comparing clones carrying a single gene copy, we show a remarkable variation in recombinant protein expression between isoforms and that the poor expression observed for one of the isoforms could only in part be explained by reduced transcript levels. Furthermore, we show that heterologous expression levels of all four aquaporin isoforms strongly respond to an increase in recombinant gene dosage, independent of the amount of protein expressed from a single gene copy. We also demonstrate that the increased expression does not appear to compromise the protein folding and the membrane localisation. Conclusions: We report a convenient and robust method based on qPCR to determine recombinant gene dosage. The method is generic for all constructs based on the pPICZ vectors and offers an inexpensive, quick and reliable means of characterising recombinant P. pastoris clones. By using this method we show that: (1) heterologous expression of all aquaporins investigated respond strongly to an increase in recombinant gene dosage (2) expression from a single recombinant gene copy varies in an isoform dependent manner (3) the poor expression observed for AtSIP1;1 is mainly caused by posttranscriptional limitations. The protein folding and membrane localisation seems to be unaffected by increased expression levels. Thus a screen for elevated gene dosage can routinely be performed for identification of P. pastoris clones with high expression levels of aquaporins and other classes of membrane proteins.
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