The mcm17 mutation of yeast shows a size-dependent segregational defect of a mini-chromosome

Abstract

Mini-chromosome-maintenance (mcm) mutants were described earlier as yeast mutants which could not stably maintain mini-chromosomes. Out of these, the ARS-specific class has been more extensively studied and is found to lose chromosomes and mini-chromosomes due to a defect in the initiation of DNA replication at yeast ARSs. In the present study we have identified a number of mcm mutants which show size-dependent loss of mini-chromosomes. When the size of the mini-chromosome was increased, from about 15 kb to about 60 kb, there was a dramatic increase in its mitotic stability in these mutants, but not in the ARS-specific class of mutants. One mutant, mcm17, belonging to the size-dependent class was further characterized. In this mutant, cells carried mini-chromosomes in significantly elevated copy numbers, suggesting a defect in segregation. This defect was largely suppressed in the 60-kb mini-chromosome. A non-centromeric plasmid, the TRP1ARS1 circle, was not affected in its maintenance. This mutant also displayed enhanced chromosome-III loss during mitosis over the wild-type strain, without elevating mitotic recombination. Cloning and sequencing of MCM17 has shown it to be the same as CHL4, a gene required for chromosome stability. This gene is non-essential for growth, as its disruption or deletion from the chromosome did not affect the growth-rate of cells at 23 degrees C or 37 degrees C. This work suggests that centromere-directed segregation of a chromosome in yeast is strongly influenced by its length.

Diversity in Requirement of Genetic and Epigenetic Factors for Centromere Function in Fungi

“…2A). A centromeric plasmid (YCp) stably propagates in the wild-type strain but exhibited very low mitotic stability when introduced into this mutant, suggesting that de novo assembly of a kinetochore on CEN DNA requires the Chl4 protein (107). Interestingly, when the same centromeric plasmid was introduced into a wild-type cell and the CHL4 gene was then deleted, two classes of transformants were obtained.…”

Ctf3p, the Mis6 budding yeast homolog, interacts with Mcm22p and Mcm16p at the yeast outer kinetochore

“…Mcm21p/Ctf5p is a member of the outer kinetochore complex that interacts with Ctf19p and Okp1p (Ortiz et al 1999). Chl4p/Ctf17p/ Mcm17p is predicted to function at the kinetochore (Kouprina et al 1993;Roy et al 1997). A different allele of ctf17 (ctf17-61) was identified previously in a CTF13 overexpression SDL screen (Kroll et al 1996).…”

“…Secondary screens applied to the ctf mutant collection successfully identified Ctf13p, Ndc10p/Ctf14p, and Ctf19p as kinetochore proteins (Doheny et al 1993;Kroll et al 1996;Hyland et al 1999). In addition to a high rate of chromosome loss, a subset of mcm, chl, and ctf mutants display phenotypes reminiscent of kinetochore mutants (Doheny et al 1993;Kouprina et al 1993;Roy et al 1997;Sanyal et al 1998;Poddar et al 1999;Ghosh et al 2001). Thus new kinetochore proteins remain to be identified from these chromosome loss mutant collections.…”

The Mcm Complex: Unwinding the Mechanism of a Replicative Helicase

“…There are also several additional genes identified in the same screen that are designated MCM but encode proteins with no homology to . These include Mcm10, another essential DNA replication factor (discussed further below), as well as others (e.g., Mcm17 [also known as Chl4] [211], Mcm21, and Mcm22 [199]) that function in chromosome segregation.…”