Chromosome condensation and sister chromatid pairing in budding yeast.

Abstract

Abstract. We have developed a fluorescent in situ hybridization (FISH) method to examine the structure of both natural chromosomes and small artificial chromosomes during the mitotic cycle of budding yeast. Our results suggest that the pairing of sister chromatids: (a) occurs near the centromere and at multiple places along the chromosome arm as has been observed in other eukaryotic cells; (b) is maintained in the absence of catenation between sister DNA molecules; and (c) is independent of large blocks of repetitive DNA commonly associated with heterochromatin. Condensation of a unique region of chromosome XVI and the highly repetitive ribosomal DNA (rDNA) cluster from chromosome XII were also examined in budding yeast. Interphase chromosomes were condensed 80-fold relative to B form DNA, similar to what has been observed in other eukaryotes, suggesting that the structure of interphase chromosomes may be conserved among eukaryotes. While additional condensation of budding yeast chromosomes were observed during mitosis, the level of condensation was less than that observed for human mitotic chromosomes. At most stages of the cell cycle, both unique and repetitive sequences were either condensed or decondensed. However, in cells arrested in late mitosis (M) by a cdc15 mutation, the unique DNA appeared decondensed while the repetitive rDNA region appeared condensed, suggesting that the condensation state of separate regions of the genome may be regulated differently. The ability to monitor the pairing and condensation of sister chromatids in budding yeast should facilitate the molecular analysis of these processes as well as provide two new landmarks for evaluating the function of important cell cycle regulators like p~ kinases and cyclins. Finally our FISH method provides a new tool to analyze centromeres, telomeres, and gene expression in budding yeast.EPLICATED chromosomes (sister chromatids) are paired and condensed prior to their segregation in mitosis. The pairing between sister chronmtids is needed to establish a stable bipolar attachment of sister chromatids to microtubules emanating from opposite poles of the mitotic spindle (Ault and Nicklas, 1989). This bipolar attachment in turn helps ensure that sister chromatids segregate from each other during armphase. In addition, the dissolution of pairing appears to be a key event in governing the onset of chromosome segregation, more commonly known as the ~meta-phase to anaphase transition: The condensation of chromosomes shortens their length which may serve to minimize the entanglement of chromosomes with one other while they are being moved by the mitotic apparatus. This shortening may also help ensure that the lagging ends of segregating chromosomes are moved far enough away from the plane of cell division so as not to be cleaved by cytokinesis. Hence, both pairing and condensation of sister chromatids appear to be essential for proper chromosome segregation.

The INO80 Chromatin Remodeling Complex Functions in Sister Chromatid Cohesion

“…21 Because sister chromatid cohesion is present from S phase through metaphase, each mitotic cell contains one fluorescent signal, whereas cells with precociously separated sisters contain two signals. 22 Cells were grown logarithmically and arrested in G 2 /M with nocoda�ole or in G 1 with a-factor. The number of GFP spots per cell was then determined (Fig.…”

In Vivo Analysis of Chromosome Condensation inSaccharomyces cerevisiae

“…Although analysis of the rDNA locus has proven to be a valuable approach, other regions of the genome have been largely neglected. Only a few studies have correlated rDNA condensation to condensation of chromosome regions that are not highly repetitive (Guacci et al, 1994;Lavoie et al, 2000). These studies observed condensation of chromosome XVI using FISH probes 145 kb and/or 255 kb apart.…”

“…However, several indirect methods to visualize chromosomes have been developed (reviewed in Loidl, 2003). Visualization of chromosome condensation using fluorescent in situ hybridization (FISH) in budding yeast revealed that highly repetitive rDNA sequences become compact during mitosis (Guacci et al, 1994) and hence this locus became the paradigm to study the process of condensation in yeast. FISH analysis of the rDNA locus revealed that this region of the genome undergoes dramatic structural changes through the cell cycle.…”

Cell cycle control of spindle elongation

“…37 The linear chromosome compaction ratio in interphase falls between 70-100 in both yeast 32 and mammalian cells. 38,39 For mitotic chromosomes this number is estimated to be 3,000-2,00,000 in vertebrates [39][40][41] but only 140 in S. cerevisiae 32 and 700 in S. pombe. 42 Thus, the degree of chromosome compaction in mitosis compared to interphase rises dramatically in higher eukaryotes.…”

“…Although human cells deficient of anaphase A are able to separate condensation in mitosis is only marginal in yeast cells and the chromosomes still occupy relatively large space in the nucleus compared to the size of the metaphase spindle. [31][32][33] Thus, depolymerization of the short kMTs is not effective in assisting chromosome segregation and separation of the chromosome arms is only achieved after the spindle poles move apart (Fig. 1B).…”