Characterization of Schizosaccharomyces pombe minichromosome deletion derivatives and a functional allocation of their centromere.

Abstract

A 530 kb long Schizosaccharomyces pombe linear minichromosome, Chl6, containing a centric region of chromosome m, has previously been made. In the present study, we constructed a number of deletions in the right and/or left arms of Chl6, and compared their structure and behaviour with Chl6. The functional centromere, cen3, is allocated within a 120 kb long region which is covered by the shortest derivative, ChlO, and is comprised mostly of centromeric repeating sequences. The shortest minichromosome is stable in mitosis and the copy number control is apparently precise. In monosomic meiosis it segregates normally. In disomic meioses, however, the frequency of non-disjunction is very high, suggesting that it may not form a pair. The mitotic loss rate of one of the left-arm deletions, ChR32, which lacks a part of the centromeric repeating sequence, is the highest of all the deletions. This deletion also exhibits the highest precocious sister chromatid separation in meiosis I, suggesting that sister chromatid association might become weakened in ChR32. Our results indicate that the proper meiotic segregation of S.pombe minichromosomes is dependent upon the formation of a bivalent. S.pombe may not have the 'distributive segregation' found with Saccharomyces cerevisiae minichromosomes.

Elimination of shelterin components bypasses RNAi for pericentric heterochromatin assembly

“…6A; Supplemental S5B). Conversely, cells containing the minichromosome Ch10 (Niwa et al 1989), which is mainly composed of pericentric repeats and centromere and telomere sequences, show reduced levels of Swi6 selectively at subtelomeric regions (Chikashige et al 2007). It is worth noting that the budding yeast, which uses Sir2/3/4 proteins to assemble heterochromatin instead of the H3K9me-HP1 system, also uses telomeres to limit silencing factors, and releasing these factors from telomeres similarly increases silencing at internal sites (Maillet et al 1996;Marcand et al 1996;Taddei et al 2009).…”

Requirement of Fission Yeast Cid14 in Polyadenylation of rRNAs

“…Consistent with these apparent chromosome segregation defects, cid14⌬ mutants had a moderate increase in the rate of minichromosome loss. Mean loss rates were 0.14% and 0.04% per division for cid14⌬ and wild-type strains, respectively (average of three independent clones), using strains containing the nonessential ade6-M216-marked Ch16 minichromosome derivative of chromosome 3 (25). Despite the chromosome segregation defects described, cid14⌬ mutants are viable, indicating that the spindle checkpoint might be required for the survival of these cells.…”

TORC2 Is Required to Maintain Genome Stability during S Phase in Fission Yeast

“…We used ⌬tor1, ⌬ste20, and ⌬gad8 strains that carried a copy of the artificial Ch16. Ch16 is a highly stable 530-kb linear minichromosome that contains a centric region of chromosome III (64). Ch16 carries an ade6-M216 point mutation that is capable of complementing an ade6-M210 mutation, thus leading to an ade ϩ phenotype.…”