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Genetic and molecular analysis of novel genes involved in chromosome segregation in Saccharomyces cerevisiae
Abstract
A genetic approach using a colony color assay to monitor the mitotic segregation of a SUP11-marked chromosome III bearing a partially functional centromere was used to identify new genes involved in chromosome segregation. Two cold sensitive (Cs) alleles, cse1-1 and cse2-1, were identified which abolish the partial function of a mutant centromere but have a little effect on the function of wildtype centromeres and exhibit no increase mitotic recombination. The cse2-1 mutant is also temperature sensitive (Ts). Both cse1-1 and cse2-1 mutants are more sensitive to the microtubule depolymerizing drug nocodazole than wildtype cells and exhibit accumulation of large-budded cells with abnormal nuclear localization and deformed spindle structures at the nonpermissive temperature. Suppressor analysis suggests that the CSE1 and CSE2 gene products may functionally interact. CSE1 and CSE2 were cloned by complementing the Cs phenotypes of cse1-1 and cse2-1 strains, respectively. CSE1 is located adjacent to the HAP2 gene on the left arm of chromosome VII and encodes a putative 110-kD protein containing 960 amino acid residues. The CSE1 protein is highly negatively charged and contains two putative phosphorylation sites and two heptad repeats of hydrophobic amino acids. Gene disruption shows that CSE1 is essential for cell growth. The CSE2 gene is located 3 cM from CEN14 on the right arm of chromosome XIV and encodes a putative 17-kD protein containing 149 amino acid residues. The CSE2 protein contains two putative phosphorylation sites and a leucine zipper motif at the carboxy terminus. CSE2 is not essential for cell viability but disruption of CSE2 results in defective mitosis and meiosis, including slower growth at 30$\sp\circ$C, cold and temperature sensitivity, increased missegregation of the X69-chromosome, accumulation of large-budded cells with abnormal nuclear and spindle morphologies at the nonpermissive temperature, reduced sporulation efficiency, formation of abnormal asci, and lower spore viability. Analysis of double and triple mutants containing cse2::LEU2, a mutant centromere and cep1::URA3 (lacking a centromere-binding protein) indicates that the CSE2 protein could be directly involved in kinetochore function. Two high dosage suppressor genes, SCM1 and SCM2, were identified which rescue the Cs phenotypes of cse1-1 and cse2-1, respectively. SCM1 is located adjacent to KAR1 on the left arm of chromosome XIV and SCM2 is located on chromosome VII. The SCM1 gene encodes a putative 60-kD protein containing 542 amino acid residues. The SCM1 protein contains a putative phosphorylation site and two heptad repeats of hydrophobic amino acid residues. Disruption of SCM1 results in cell inviability indicating that SCM1 is essential for cell growth. SCM2 is also a novel gene indispensable for cell viability. In conclusion, I have discovered four novel genes, CSE1, CSE2, SCM1, and SCM2 which are involved in chromosome segregation and encode gene products that are functionally related and may physically interact.
Subject Area
Genetics|Molecular biology
Recommended Citation
Xiao, Zhixiong, "Genetic and molecular analysis of novel genes involved in chromosome segregation in Saccharomyces cerevisiae" (1991). Doctoral Dissertations Available from Proquest. AAI9207475.
https://scholarworks.umass.edu/dissertations/AAI9207475