The alleles cse1-1 and cse2-l were cold-sensitive yeast mutants that display increased mitotic chromosome nondisjunction phenotypes. Two high dosage suppressors, SCM1 and SCM2, were isolated which suppress the cold-sensitive phenotypes of cse1-1 and cse2-1 cells, respectively. DNA sequence analysis revealed that SCM1 is identical to SRP1, which was previously isolated as an extragenic suppressor of an RNA polymerase I mutant. SRP1 encodes a 67 kD protein containing an arm (armadillo) motif that localizes to the periphery of the nucleus. Mutational analysis indicates that SRP1 is essential for viability and that the C-terminus is important for function. Genetic analysis indicates that SRP1 and CSE1 functionally interact in chromosome transmission. In addition, a putative mouse homolog of SRP1 called MSG1 was isolated, which also contains an arm motif in the central domain. Overexpression of MSG1 suppresses the cold-sensitive phenotype of cse1-1 cells suggesting a possible functional similarity between SRP1 and MSG1. In addition to the cold-sensitive and chromosome missegregation phenotypes, cse2-1 also causes temperature sensitivity. Overexpression of SCM2 only suppresses the cold-sensitive phenotype. SCM2 encodes a 65 kD protein that is highly homologous to known amino acid permeases. Disruption of SCM2 causes slow growth. The scm2$\Delta$ trp1-$\Delta$101 double mutant cells exhibit a synthetic cold-sensitive phenotype and grow much more slowly at the permissive temperature than cells with either a scm2$\Delta$ or trp1-$\Delta$101 single mutation. Amino acid uptake assays revealed a decreased accumulation of tryptophan in the scm$\Delta$ cells indicating that SCM2 encodes a tryptophan permease. A temperature-sensitive allele of phosphoinositide-specific phospholipase C gene in yeast, plc1-1, causes a 9-fold increase in chromosome missegregation. The temperature sensitivity and the chromosome missegregation phenotypes of plc1-1 cells were both partially suppressed by calcium. The mutant plc1-1 allele was cloned and found to encode a single amino acid substitution in the protein. The yeast PLC1 protein contains a putative EF-hand calcium binding domain and the enzymatic activity of PLC1 is dependent upon calcium. Five EF-hand mutants have been generated. Three of these mutants exhibit a temperature-sensitive phenotype that can be suppressed by calcium. The effects of the plc1 mutations on chromosome segregation and cell lysis were also investigated.
| Identifer | oai:union.ndltd.org:UMASS/oai:scholarworks.umass.edu:dissertations-9080 |
| Date | 01 January 1995 |
| Creators | Chen, Xiao Hong |
| Publisher | ScholarWorks@UMass Amherst |
| Source Sets | University of Massachusetts, Amherst |
| Language | English |
| Detected Language | English |
| Type | text |
| Source | Doctoral Dissertations Available from Proquest |
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