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Accurate chromosome segregation is necessary to ensure genomic integrity. Segregation depends on the proper functioning of the centromere, kinetochore, and mitotic spindle microtubules and is monitored by the spindle assembly checkpoint (SAC). In the fission yeast Schizosaccharomyces pombe, defects in Dis1, a microtubule-associated protein that influences microtubule dynamics, lead to mitotic arrest as a result of an active SAC and consequent failure to grow at low temperature. In a mutant dis1 background (dis1-288), loss of function of Msc1, a fission yeast homolog of the KDM5 family of proteins, suppresses the growth defect and promotes normal mitosis. Genetic analysis implicates a histone deacetylase (HDAC)–linked pathway in suppression because HDAC mutants clr6-1, clr3∆, and sir2∆, though not hos2∆, also promote normal mitosis in the dis1-288 mutant. Suppression of the dis phenotype through loss of msc1 function requires the spindle checkpoint protein Mad2 and is limited by the presence of the heterochromatin-associated HP1 protein homolog Swi6. We speculate that alterations in histone acetylation promote a centromeric chromatin environment that compensates for compromised dis1 function by allowing for successful kinetochore-microtubule interactions that can satisfy the SAC. In cells arrested in mitosis by mutation of dis1, loss of function of epigenetic determinants such as Msc1 or specific HDACs can promote cell survival. Because the KDM5 family of proteins has been implicated in human cancers, an appreciation of the potential role of this family of proteins in chromosome segregation is warranted. 相似文献
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Mahnaz Amini Yiming Chang Ulrich Wissenbach Veit Flockerzi Gabriel Schlenstedt Andreas Beck 《The Journal of biological chemistry》2021,297(4)
Transient receptor potential (TRP) cation channels, which are conserved across mammals, flies, fish, sea squirts, worms, and fungi, essentially contribute to cellular Ca2+ signaling. The activity of the unique TRP channel in yeast, TRP yeast channel 1 (TRPY1), relies on the vacuolar and cytoplasmic Ca2+ concentration. However, the mechanism(s) of Ca2+-dependent regulation of TRPY1 and possible contribution(s) of Ca2+-binding proteins are yet not well understood. Our results demonstrate a Ca2+-dependent binding of yeast calmodulin (CaM) to TRPY1. TRPY1 activity was increased in the cmd1–6 yeast strain, carrying a non–Ca2+-binding CaM mutant, compared with the parent strain expressing wt CaM (Cmd1). Expression of Cmd1 in cmd1–6 yeast rescued the wt phenotype. In addition, in human embryonic kidney 293 cells, hypertonic shock-induced TRPY1-dependent Ca2+ influx and Ca2+ release were increased by the CaM antagonist ophiobolin A. We found that coexpression of mammalian CaM impeded the activity of TRPY1 by reinforcing effects of endogenous CaM. Finally, inhibition of TRPY1 by Ca2+–CaM required the cytoplasmic amino acid stretch E33–Y92. In summary, our results show that TRPY1 is under inhibitory control of Ca2+–CaM and that mammalian CaM can replace yeast CaM for this inhibition. These findings add TRPY1 to the innumerable cellular proteins, which include a variety of ion channels, that use CaM as a constitutive or dissociable Ca2+-sensing subunit, and contribute to a better understanding of the modulatory mechanisms of Ca2+–CaM. 相似文献
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Expression Levels of the Yeast Alcohol Acetyltransferase Genes ATF1, Lg-ATF1, and ATF2 Control the Formation of a Broad Range of Volatile Esters 总被引:3,自引:0,他引:3
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Kevin J. Verstrepen Stijn D. M. Van Laere Bart M. P. Vanderhaegen Guy Derdelinckx Jean-Pierre Dufour Isak S. Pretorius Joris Winderickx Johan M. Thevelein Freddy R. Delvaux 《Applied microbiology》2003,69(9):5228-5237
Volatile aroma-active esters are responsible for the fruity character of fermented alcoholic beverages such as beer and wine. Esters are produced by fermenting yeast cells in an enzyme-catalyzed intracellular reaction. In order to investigate and compare the roles of the known Saccharomyces cerevisiae alcohol acetyltransferases, Atf1p, Atf2p and Lg-Atf1p, in volatile ester production, the respective genes were either deleted or overexpressed in a laboratory strain and a commercial brewing strain. Subsequently, the ester formation of the transformants was monitored by headspace gas chromatography and gas chromatography combined with mass spectroscopy (GC-MS). Analysis of the fermentation products confirmed that the expression levels of ATF1 and ATF2 greatly affect the production of ethyl acetate and isoamyl acetate. GC-MS analysis revealed that Atf1p and Atf2p are also responsible for the formation of a broad range of less volatile esters, such as propyl acetate, isobutyl acetate, pentyl acetate, hexyl acetate, heptyl acetate, octyl acetate, and phenyl ethyl acetate. With respect to the esters analyzed in this study, Atf2p seemed to play only a minor role compared to Atf1p. The atf1Δ atf2Δ double deletion strain did not form any isoamyl acetate, showing that together, Atf1p and Atf2p are responsible for the total cellular isoamyl alcohol acetyltransferase activity. However, the double deletion strain still produced considerable amounts of certain other esters, such as ethyl acetate (50% of the wild-type strain), propyl acetate (50%), and isobutyl acetate (40%), which provides evidence for the existence of additional, as-yet-unknown ester synthases in the yeast proteome. Interestingly, overexpression of different alleles of ATF1 and ATF2 led to different ester production rates, indicating that differences in the aroma profiles of yeast strains may be partially due to mutations in their ATF genes. 相似文献
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Sophie D Lefevre Sanjeev Kumar Ida J van der Klei 《Cell cycle (Georgetown, Tex.)》2015,14(11):1698-1703
Mitochondria are key players in aging and cell death. It has been suggested that mitochondrial fragmentation, mediated by the Dnm1/Fis1 organelle fission machinery, stimulates aging and cell death. This was based on the observation that Saccharomyces cerevisiae Δdnm1 and Δfis1 mutants show an enhanced lifespan and increased resistance to cell death inducers. However, the Dnm1/Fis1 fission machinery is also required for peroxisome division. Here we analyzed the significance of peroxisome fission in yeast chronological lifespan, using yeast strains in which fission of mitochondria was selectively blocked. Our data indicate that the lifespan extension caused by deletion of FIS1 is mainly due to a defect in peroxisome fission and not caused by a block in mitochondrial fragmentation. These observations are underlined by our observation that deletion of FIS1 does not lead to lifespan extension in yeast peroxisome deficient mutant cells. 相似文献
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Eugenol has antifungal activity and is recognised as having therapeutic potential. However, little is known of the cellular basis of its antifungal activity and a better understanding of eugenol tolerance should lead to better exploitation of eugenol in antifungal therapies. The model yeast, Saccharomyces cerevisiae, expressing apoaequorin was used to show that eugenol induces cytosolic Ca2+ elevations. We investigated the eugenol Ca2+ signature in further detail and show that exponentially growing cells exhibit Ca2+ elevation resulting exclusively from the influx of Ca2+ across the plasma membrane whereas in stationary growth phase cells Ca2+ influx from intracellular and extracellular sources contribute to the eugenol-induced Ca2+ elevation. Ca2+ channel deletion yeast mutants were used to identify the pathways mediating Ca2+ influx; intracellular Ca2+ release was mediated by the vacuolar Ca2+ channel, Yvc1p, whereas the Ca2+ influx across the plasma membrane could be resolved into Cch1p-dependent and Cch1p-independent pathways. We show that the growth of yeast devoid the plasma membrane Ca2+ channel, Cch1p, was hypersensitive to eugenol and that this correlated with reduced Ca2+ elevations. Taken together, these results indicate that a cch1p-mediated Ca2+ influx is part of an intracellular signal which protects against eugenol toxicity. This study provides fresh insight into the mechanisms employed by fungi to tolerate eugenol toxicity which should lead to better exploitation of eugenol in antifungal therapies. 相似文献
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Sha Wang Jinling Cao Xiao Liu Hongqin Hu Jie Shi Shizhu Zhang Nancy P. Keller Ling Lu 《PloS one》2012,7(10)
Although the high affinity Ca2+ channel, Cch1, and its subunit Mid1 have been investigated and evaluated in yeast and some of filamentous fungi, little is known about the function of their homologs in the Aspergilli. Here, we have functionally characterized the yeast homologs, CchA and MidA, in Aspergillus nidulans using conditional and null deletion mutants. CchA and MidA not only have functional benefits of fast growth, which is consistent with Cch1 and Mid1 in yeast, but also have unique and complex roles in regulating conidiation, hyphal polarity and cell wall components in low-calcium environments. The defect of CchA or MidA resulted in a sharp reduction in the number of conidiospores, accompanied by abnormal metulae, and undeveloped-phialides at a higher density of inoculum. Most interestingly, these conidiation defects in mutants can, remarkably, be rescued either by extra-cellular Ca2+ in a calcineurin-dependent way or by osmotic stress in a calcineurin-independent way. Moreover, the fact that the phenotypic defects are not exacerbated by the presence of the double deletion, together with the Y2H assay, indicates that CchA and MidA may form a complex to function together. Our findings suggest that the high-affinity Ca2+ channel may represent a viable and completely unexplored avenue to reduce conidiation in the Aspergilli. 相似文献
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Mitochondria are dynamic organelles that constantly undergo fission, fusion, and movement. Increasing evidence indicates that these dynamic changes are intricately related to mitochondrial function, suggesting that mitochondrial form and function are linked. Calcium (Ca2+) is one signal that has been shown to both regulate mitochondrial fission in various cell types and stimulate mitochondrial enzymes involved in ATP generation. However, although Ca2+ plays an important role in adult cardiac muscle cells for excitation–metabolism coupling, little is known about whether Ca2+ can regulate their mitochondrial morphology. Therefore, we tested the role of Ca2+ in regulating cardiac mitochondrial fission. We found that neonatal and adult cardiomyocyte mitochondria undergo rapid and transient fragmentation upon a thapsigargin (TG)- or KCl-induced cytosolic Ca2+ increase. The mitochondrial fission protein, DLP1, participates in this mitochondrial fragmentation, suggesting that cardiac mitochondrial fission machinery may be regulated by intracellular Ca2+ signaling. Moreover, the TG-induced fragmentation was also associated with an increase in reactive oxygen species (ROS) formation, suggesting that activation of mitochondrial fission machinery is an early event for Ca2+-mediated ROS generation in cardiac myocytes. These results suggest that Ca2+, an important regulator of muscle contraction and energy generation, also dynamically regulates mitochondrial morphology and ROS generation in cardiac myocytes. 相似文献
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Michela Carraro Valentina Giorgio Justina ?ileikyt? Geppo Sartori Michael Forte Giovanna Lippe Mario Zoratti Ildikò Szabò Paolo Bernardi 《The Journal of biological chemistry》2014,289(23):15980-15985
Purified F-ATP synthase dimers of yeast mitochondria display Ca2+-dependent channel activity with properties resembling those of the permeability transition pore (PTP) of mammals. After treatment with the Ca2+ ionophore ETH129, which allows electrophoretic Ca2+ uptake, isolated yeast mitochondria undergo inner membrane permeabilization due to PTP opening. Yeast mutant strains ΔTIM11 and ΔATP20 (lacking the e and g F-ATP synthase subunits, respectively, which are necessary for dimer formation) display a striking resistance to PTP opening. These results show that the yeast PTP originates from F-ATP synthase and indicate that dimerization is required for pore formation in situ. 相似文献
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Emilie Castonguay Sharon A. White Alexander Kagansky Daniel J. St-Cyr Araceli G. Castillo Christiane Brugger Rachel White Carolina Bonilla Michaela Spitzer William C. Earnshaw Thomas Schalch Karl Ekwall Mike Tyers Robin C. Allshire 《Molecular and cellular biology》2015,35(4):662-674
Heterochromatin underpins gene repression, genome integrity, and chromosome segregation. In the fission yeast Schizosaccharomyces pombe, conserved protein complexes effect heterochromatin formation via RNA interference-mediated recruitment of a histone H3 lysine 9 methyltransferase to cognate chromatin regions. To identify small molecules that inhibit heterochromatin formation, we performed an in vivo screen for loss of silencing of a dominant selectable kanMX reporter gene embedded within fission yeast centromeric heterochromatin. Two structurally unrelated compounds, HMS-I1 and HMS-I2, alleviated kanMX silencing and decreased repressive H3K9 methylation levels at the transgene. The decrease in methylation caused by HMS-I1 and HMS-I2 was observed at all loci regulated by histone methylation, including centromeric repeats, telomeric regions, and the mating-type locus, consistent with inhibition of the histone deacetylases (HDACs) Clr3 and/or Sir2. Chemical-genetic epistasis and expression profiles revealed that both compounds affect the activity of the Clr3-containing Snf2/HDAC repressor complex (SHREC). In vitro HDAC assays revealed that HMS-I1 and HMS-I2 inhibit Clr3 HDAC activity. HMS-I1 also alleviated transgene reporter silencing by heterochromatin in Arabidopsis and a mouse cell line, suggesting a conserved mechanism of action. HMS-I1 and HMS-I2 bear no resemblance to known inhibitors of chromatin-based activities and thus represent novel chemical probes for heterochromatin formation and function. 相似文献