Genome-wide haploinsufficiency screen reveals a novel role for γ-TuSC in spindle organization and genome stability

Choy, John S.; O’Toole, Eileen; Schuster, Breanna M.; Crisp, Matthew J.; Karpova, Tatiana S.; McNally, James G.; Winey, Mark; Gardner, Melissa K.; Basrai, Munira A.

doi:10.1091/mbc.e12-12-0902

Cited by 11 publications

(11 citation statements)

References 62 publications

(33 reference statements)

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“…We observed that all three high-copy TPKs resulted in 6to 12-fold increase in chromosome loss when compared to empty vector control (Table 1). This result suggests that increased PKA activity leads to greater chromosome instability, consistent with our previous report showing that diploid yeast with a hemizygous deletion of BCY1 have increased rates of chromosome loss compared to wild-type cells (Magtanong et al 2011;Choy et al 2013). Together, these observations indicate that PKA antagonizes Dam1p activity and increases errors in chromosome segregation.…”

Section: High-copy Tpk1 Tpk2 and Tpk3 Antagonize Dam1p Function Andsupporting

confidence: 91%

Glucose Signaling Is Connected to Chromosome Segregation Through Protein Kinase A Phosphorylation of the Dam1 Kinetochore Subunit inSaccharomyces cerevisiae

et al. 2018

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The Dam1 complex is an essential component of the outer kinetochore that mediates attachments between spindle microtubules and chromosomes. Dam1p, a subunit of the Dam1 complex, binds to microtubules and is regulated by Aurora B/Ipl1p phosphorylation. We find that overexpression of cAMP-dependent protein kinase (PKA) catalytic subunits (i.e., TPK1, TPK2, TPK3) is lethal in DAM1 mutants and increases the rate of chromosome loss in wild-type cells. Replacing an evolutionarily conserved PKA site (S31) in Dam1p with a nonphosphorylatable alanine suppressed the high-copy PKA dosage lethality in dam1-1. Consistent with Dam1p as a target of PKA, we find that in vitro PKA can directly phosphorylate S31 in Dam1p and we observed phosphorylation of S31 in Dam1p purified from asynchronously growing yeast cells. Cells carrying high-copy TPK2 or a Dam1p phospho-mimetic S31D mutant displayed a reduction in Dam1p localization at the kinetochore, suggesting that PKA phosphorylation plays a role in assembly and/or stability of the Dam1 complex. Furthermore, we observed spindle defects associated with S31 phosphorylation. Finally, we find that phosphorylation of Dam1p on S31 is reduced when glucose is limiting as well as during a-factor arrest, conditions that inhibit PKA activity. These observations suggest that the PKA site of Dam1p participates in regulating kinetochore activity. While PKA is a wellestablished effector of glucose signaling, our work shows for the first time that glucose-dependent PKA activity has an important function in chromosome segregation.

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Section: High-copy Tpk1 Tpk2 and Tpk3 Antagonize Dam1p Function Andsupporting

confidence: 91%

Glucose Signaling Is Connected to Chromosome Segregation Through Protein Kinase A Phosphorylation of the Dam1 Kinetochore Subunit inSaccharomyces cerevisiae

et al. 2018

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“…However, unlike strong loss of function mutations gene dosage effects are likely to be relatively subtle and requiring reliable and sensitive quantitative assays. Two previous studies ( Choy et al 2013 ; Strome et al 2008 ) identified sets of genes that cause increased CIN when hemizygous. We refer to these genes as dcCIN genes.…”

Section: Resultsmentioning

confidence: 99%

Single-Cell Based Quantitative Assay of Chromosome Transmission Fidelity

Zhu

Heinecke

Mulla

et al. 2015

G3 Genes|Genomes|Genetics

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Errors in mitosis are a primary cause of chromosome instability (CIN), generating aneuploid progeny cells. Whereas a variety of factors can influence CIN, under most conditions mitotic errors are rare events that have been difficult to measure accurately. Here we report a green fluorescent protein−based quantitative chromosome transmission fidelity (qCTF) assay in budding yeast that allows sensitive and quantitative detection of CIN and can be easily adapted to high-throughput analysis. Using the qCTF assay, we performed genome-wide quantitative profiling of genes that affect CIN in a dosage-dependent manner and identified genes that elevate CIN when either increased (icCIN) or decreased in copy number (dcCIN). Unexpectedly, qCTF screening also revealed genes whose change in copy number quantitatively suppress CIN, suggesting that the basal error rate of the wild-type genome is not minimized, but rather, may have evolved toward an optimal level that balances both stability and low-level karyotype variation for evolutionary adaptation.

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“…This is likely an underestimation, since many genes may be haploinsufficient in conditions that are not tested. Indeed, a study that assessed genome instability reported additional haploinsufficient genes 24 . Haploinsufficient genes are enriched in genome ontology categories relating to transcription and transcript processing, protein folding, protein transport, and ribosome biogenesis 23 .…”

Section: Why Chromosome Dosage Mattersmentioning

confidence: 99%

Mechanisms of X Chromosome Dosage Compensation

Ercan

2015

J. Genomics

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In many animals, males have one X and females have two X chromosomes. The difference in X chromosome dosage between the two sexes is compensated by mechanisms that regulate X chromosome transcription. Recent advances in genomic techniques have provided new insights into the molecular mechanisms of X chromosome dosage compensation. In this review, I summarize our current understanding of dosage imbalance in general, and then review the molecular mechanisms of X chromosome dosage compensation with an emphasis on the parallels and differences between the three well-studied model systems, M. musculus, D. melanogaster and C. elegans.

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Genome-wide haploinsufficiency screen reveals a novel role for γ-TuSC in spindle organization and genome stability

Cited by 11 publications

References 62 publications

Glucose Signaling Is Connected to Chromosome Segregation Through Protein Kinase A Phosphorylation of the Dam1 Kinetochore Subunit inSaccharomyces cerevisiae

Glucose Signaling Is Connected to Chromosome Segregation Through Protein Kinase A Phosphorylation of the Dam1 Kinetochore Subunit inSaccharomyces cerevisiae

Single-Cell Based Quantitative Assay of Chromosome Transmission Fidelity

Mechanisms of X Chromosome Dosage Compensation

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