| 1. |
- Alvarado-Kristensson, Maria, et al.
(författare)
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SADB phosphorylation of gamma-tubulin regulates centrosome duplication.
- 2009
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Ingår i: Nature Cell Biology. - : Springer Science and Business Media LLC. - 1465-7392 .- 1476-4679. ; 11:9, s. 86-1081
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Tidskriftsartikel (refereegranskat)abstract
- Symmetrical cell division requires duplication of DNA and protein content to generate two daughter cells. Centrosomes also duplicate during cell division, but the mechanism controlling this process is incompletely understood. We describe an alternative splice form of SadB encoding a short SADB Ser/Thr kinase whose activity fluctuates during the cell cycle, localizes to centrosomes, and controls centrosome duplication. Reduction of endogenous SADB levels diminished centrosome numbers, whereas enhanced SADB expression induced centrosome amplification. SADB exerted this action through phosphorylation of gamma-tubulin on Ser 131, as expression of a phosphomimetic Ser 131-to-Asp gamma-tubulin mutant alone increased centrosome numbers, whereas non-phosphorylatable Ala 131-gamma-tubulin impaired centrosome duplication. We propose that SADB kinase activity controls centrosome homeostasis by regulating phosphorylation of gamma-tubulin.
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| 2. |
- Alvarado-Kristensson, Maria
(författare)
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A simple and fast method for fixation of cultured cell lines that preserves cellular structures containing gamma-tubulin
- 2018
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Ingår i: MethodsX. - : Elsevier BV. - 2215-0161. ; 5, s. 227-233
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Tidskriftsartikel (refereegranskat)abstract
- When using fluorescence microscope techniques to study cells, it is essential that the cell structure and contents are preserved after preparation of the samples, and that the preparation method employed does not create artefacts that can be perceived as cellular structure/components. γ-Tubulin forms filaments that in some cases are immunostained with an anti-γ-tubulin antibody, but this immunostaining is not reproducible [1,2]. In addition, the C terminal region of γ-tubulin (green fluorescence protein tagged [GFP]-γ-tubulin334––449) forms cytosolic GFP-labeled structures, which can easily be imaged in live cells but are not preserved in fixed cells [1,3]. The purpose of this study was to identify a fixation technique that preserves cellular constituents containing γ-tubulin. • This protocol describes a method that preserves γ-tubulin-containing structures in fixed cells.• The technique entails two-step fixation. A pre-fixation step using paraformaldehyde is followed by a final fixation and permeabilization step performed at −80 °C.• In comparison with other methodology for fixation [4–6], the technique presented here uses a short pre-fixation step with a mixture of paraformaldehyde and sucrose followed by a short fixation/permeabilization step with a mixture of methanol and acetone at −80 °C.
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| 3. |
- Alvarado Kristensson, Maria
(författare)
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Choreography of the centrosome
- 2020
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Ingår i: Heliyon. - : Elsevier BV. - 2405-8440. ; 6:e03228, s. 1-6
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Forskningsöversikt (refereegranskat)abstract
- More than a century ago, the centrosome was discovered and described as “the true division organ of the cell”. Electron microscopy revealed that a centrosome is an amorphous structure or pericentriolar protein matrix that surrounds a pair of well-organized centrioles. Today, the importance of the centrosome as a microtubule-organizing center and coordinator of the mitotic spindle is questioned, because centrioles are absent in up to half of all known eukaryotic species, and various mechanisms for acentrosomal microtubule nucleation have been described. This review recapitulates the known functions of centrosome movements in cellular homeostasis and discusses knowledge gaps in this field.
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| 4. |
- Alvarado-Kristensson, Maria
(författare)
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Ett nytt cellskelett i kampen mot cancer
- 2018
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Ingår i: Onkologi i Sverige : den oberoende tidningen för svensk cancervård. - 1653-1582. ; 5, s. 44-48
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Forskningsöversikt (övrigt vetenskapligt/konstnärligt)abstract
- Forskare vid Lunds universitet har upptäckt ett cellskelett som ger struktur åt mito-kondrierna, cellens energifabriker. Skelettet är nödvändigt för mitokondriernas funktion, men forskarna fann även att cancerceller trots defekter utnyttjar skelettet för att kunna bibehålla sin energiproduktion – och därigenom överleva. Cellskelettet i mitokondrierna fungerar ungefär som ett armeringsnät och hjälper till att bibehålla cellens uppbyggnad och bidrar till stabiliteten i cellens olika rum. Det gör att olika molekyler lättare kan ta sig runt i cellen, skriver Maria Alvarado Kristensson, docent och forskare i molekylär patologi vid Lunds universitet.
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| 5. |
- Alvarado-Kristensson, Maria, et al.
(författare)
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p38-MAPK signals survival by phosphorylation of caspase-8 and caspase-3 in human neutrophils
- 2004
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Ingår i: Journal of Experimental Medicine. - : Rockefeller University Press. - 0022-1007 .- 1540-9538. ; 199:4, s. 449-458
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Tidskriftsartikel (refereegranskat)abstract
- Neutrophil apoptosis occurs both in the bloodstream and in the tissue and is considered essential for the resolution of an inflammatory process. Here, we show that p38-mitogen-activated protein kinase (MAPK) associates to caspase-8 and caspase-3 during neutrophil apoptosis and that p38-MAPK activity, previously shown to be a survival signal in these primary cells, correlates with the levels of caspase-8 and caspase-3 phosphorylation. In in vitro experiments, immunoprecipitated active p38-MAPK phosphorylated and inhibited the activity of the active p20 subunits of caspase-8 and caspase-3. Phosphopeptide mapping revealed that these phosphorylations occurred on serine-364 and serine-150, respectively. Introduction of mutated (S150A), but not wild-type, TAT-tagged caspase-3 into primary neutrophils made the Fas-induced apoptotic response insensitive to p38-MAPK inhibition. Consequently, p38-MAPK can directly phosphorylate and inhibit the activities of caspase-8 and caspase-3 and thereby hinder neutrophil apoptosis, and, in so doing, regulate the inflammatory response.
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| 6. |
- Alvarado-Kristensson, Maria, et al.
(författare)
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p38 Mitogen-activated protein kinase and phosphatidylinositol 3-kinase activities have opposite effects on human neutrophil apoptosis.
- 2002
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Ingår i: FASEB Journal. - : Wiley. - 1530-6860 .- 0892-6638. ; 16:1, s. 31-129
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Tidskriftsartikel (refereegranskat)abstract
- Neutrophil apoptosis is essential for resolution of inflammatory reactions. Here, we studied the role of two apoptosis/survival-associated protein kinases in this process. We discovered a previously undetected early and transient inhibition of the activity of p38 mitogen-activated protein kinase (p38 MAPK) during both spontaneous and Fas-induced apoptosis. Pharmacological inhibition of this enzyme augmented the activation of caspases and the apoptotic response, which suggests that the p38 MAPK signals survival in neutrophils. Our finding that caspase-3 activity was initiated during the transient inhibition of p38 MAPK suggests that apoptosis is initiated during this inhibition. Furthermore, such transient inhibition was counteracted by granulocyte-macrophage colony-stimulating factor, which elicits survival. We also found that neither this inhibition of p38 MAPK nor the spontaneous apoptotic response depended on Fas. Instead, the early inhibition of p38 MAPK concurred with a Fas-induced activation of phosphatidylinositol 3-kinase, inhibition of which reduced apoptosis. Thus, the Fas-induced augmentation of spontaneous apoptosis can be explained by its activation of phosphatidylinositol 3-kinase. We conclude that p38 MAPK activity represents a survival signal that is inactivated transiently during both spontaneous and Fas-induced apoptosis, whereas Fas-induced phosphatidylinositol 3-kinase activity is a proapoptotic signal in isolated human neutrophils.
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| 7. |
- Alvarado-Kristensson, Maria, et al.
(författare)
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Protein phosphatase 2A regulates apoptosis in neutrophils by dephosphorylating both p38 MAPK and its substrate caspase 3.
- 2005
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Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 280:7, s. 6238-6244
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Tidskriftsartikel (refereegranskat)abstract
- The induction of apoptosis in neutrophils is an essential event in the resolution of an inflammatory process. We found recently that the reduction of the activity of the neutrophil survival factor p38 MAPK and dephosphorylation and thus activation of caspases must occur to initiate such cell death in these leukocytes. Here, we report a previously undetected early and transient activation of protein phosphatase 2A WPM in neutrophils undergoing apoptosis. The pharmacological inhibition of this phosphatase during Fas-induced apoptosis augmented the levels of phosphorylation of both p38 MAPK and caspase 3, resulting in a decreased activity of caspase 3 and an increased neutrophil survival. The complementary finding of a time-dependent association among PP2A, p38 MAPK, and caspase 3 in intact neutrophils indicated that there is a direct regulatory link among these signaling enzymes during Fas-provoked apoptosis. Moreover, immunoprecipitated active p38 MAPK and recombinant phosphorylated caspase 3 were dephosphorylated by exposure to purified PP2A in vitro. Consequently, the early and temporary activation of PP2A in neutrophils impaired not only the p38 MAPK-mediated inhibition of caspase 3 but also restored the activity to caspase 3 that had already been phosphorylated and thereby inactivated. These findings indicate that PP2A plays a pivotal dual role in the induction of neutrophil apoptosis and therefore also in the resolution of inflammation.
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| 8. |
- Alvarado-Kristensson, Maria
(författare)
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Regulation of neutrophil apoptosis
- 2004
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The human neutrophil is the most abundant granulocyte and the major type of cell involved in an acute inflammatory response. Neutrophils are armed with various systems of enzymes, that can find and kill pathogens, but unfortunately, these "weapons" cannot distinguish between the host tissues and the "invaders." Therefore, an extensive neutrophil reaction leads to continuous release of toxic metabolites, which causes successive self-destruction of host tissues and possibly also organ failure. Such a series of destructive events has been implicated in diseases such as rheumatoid arthritis, myocardial infarction/reperfusion injury, atherogenesis, asthma, cystic fibrosis, emphysema, and vasculitis. Resolution of an acute inflammatory process depends on termination of neutrophil emigration from blood vessels and clearance of extravasated neutrophils and their metabolic products. Outside the blood vessels, neutrophils spontaneously undergo apoptosis, and are therefore removed by phagocytic cells at the site of inflammation. Neutrophil apoptosis can be modulated by several factors in the local environment, such as the Fas ligand (FasL), but the molecular mechanisms involved are poorly understood. In this dissertation thesis, I describe and elucidate intracellular signalling mechanisms that are involved in regulation of spontaneous and Fas-induced apoptosis in human neutrophils. Using two different methods it was possible to detect constitutive activity of p38 mitogen-activated protein kinase (p38) in newly isolated neutrophils. The p38 survival signal was transiently lost during both spontaneous and Fas-induced apoptosis, favoured induction of the apoptotic process. During the transient loss of p38 activity there was a temporary Fas-induced increase in phosphatidylinositol 3-kinase (PI3K) activity, which also had a pro-apoptotic impact on the neutrophils. In addition, my experiments showed that the active form of p38 associates with caspase 8 and caspase 3, which is necessary for p38-induced phosphorylation of serine-362 and serine-150 on these caspases. These biochemical modifications impair the activities, and possibly also the stability, of caspase 8 and 3 and thereby weaken the capacity of these enzymes to induce apoptosis. The results in this dissertation also demonstrate that the protein phosphatase type 2A (PP2A) can directly and independently decrease the phosphorylation levels of both p38 and caspase 3. Consequently, PP2A can increase the activity of caspase 3 by dual mechanisms and thereby promote the apoptotic response in human neutrophils.
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| 9. |
- Alvarado Kristensson, Maria, et al.
(författare)
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The Biology of the Nuclear Envelope and Its Implications in Cancer Biology
- 2019
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1422-0067. ; 20:10
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Forskningsöversikt (refereegranskat)abstract
- The formation of the nuclear envelope and the subsequent compartmentalization of the genome is a defining feature of eukaryotes. Traditionally, the nuclear envelope was purely viewed as a physical barrier to preserve genetic material in eukaryotic cells. However, in the last few decades, it has been revealed to be a critical cellular component in controlling gene expression and has been implicated in several human diseases. In cancer, the relevance of the cell nucleus was first reported in the mid-1800s when an altered nuclear morphology was observed in tumor cells. This review aims to give a current and comprehensive view of the role of the nuclear envelope on cancer first by recapitulating the changes of the nuclear envelope during cell division, second, by reviewing the role of the nuclear envelope in cell cycle regulation, signaling, and the regulation of the genome, and finally, by addressing the nuclear envelope link to cell migration and metastasis and its use in cancer prognosis.
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| 10. |
- Alvarado Kristensson, Maria
(författare)
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The Game of Tubulins
- 2021
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Ingår i: Cells. - : MDPI AG. - 2073-4409. ; 10
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Forskningsöversikt (refereegranskat)abstract
- Members of the tubulin superfamily are GTPases; the activities of GTPases are necessary for life. The members of the tubulin superfamily are the constituents of the microtubules and the γ-tubulin meshwork. Mutations in members of the tubulin superfamily are involved in developmental brain disorders, and tubulin activities are the target for various chemotherapies. The intricate functions (game) of tubulins depend on the activities of the GTP-binding domain of α-, β-, and γ-tubulin. This review compares the GTP-binding domains of γ-tubulin, α-tubulin, and β-tubulin and, based on their similarities, recapitulates the known functions and the impact of the γ-tubulin GTP-binding domain in the regulation of the γ-tubulin meshwork and cellular homeostasis.
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