Homolog pairing and sister chromatid cohesion in heterochromatin in Drosophila male meiosis I

Abstract

Drosophila males undergo meiosis without recombination or chiasmata but homologous chromosomes pair and disjoin regularly. The X-Y pair utilizes a specific repeated sequence within the heterochromatic ribosomal DNA blocks as a pairing site. No pairing sites have yet been identified for the autosomes. To search for such sites, we utilized probes targeting specific heterochromatic regions to assay heterochromatin pairing sequences and behavior in meiosis by fluorescence in situ hybridization (FISH). We found that the small fourth chromosome pairs at heterochromatic region 61 and associates with the X chromosome throughout prophase I. Homolog pairing of the fourth chromosome is disrupted when the homolog conjunction complex is perturbed by mutations in SNM or MNM. On the other hand, six tested heterochromatic regions of the major autosomes proved to be largely unpaired after early prophase I, suggesting that stable homolog pairing sites do not exist in heterochromatin of the major autosomes. Furthermore, FISH analysis revealed two distinct patterns of sister chromatid cohesion in heterochromatin: regions with stable cohesion and regions lacking cohesion. This suggests that meiotic sister chromatid cohesion is incomplete within heterochromatin and may occur at specific preferential sites.

Homologous pairing and the role of pairing centers in meiosis

“…A recent fluorescent in situ hybridization (FISH) analysis, using probes to chromosome-specific repeated sequences, failed to detect any such stable pairing sites (i.e. sites that remain paired throughout meiosis I) in the heterochromatic regions of the major autosomes (Tsai et al, 2011), thus providing further evidence against PC-based pairing of the major autosomes. However, the small fourth chromosomes did remain paired at a specific heterochromatic site in >90% of spermatocytes throughout prophase I, suggesting that the fourth chromosome contains a PC.…”

“…Furthermore, recruitment of MNM, and perhaps SNM, to autosomes depends upon the Teflon (TEF) protein, which is required for segregation of the autosomes but not the sex chromosomes (Tomkiel et al, 2001;Thomas et al, 2005). We have suggested, by analogy to chiasmata in recombinational meiosis, that stable connections between autosomal homologs exist at different sites in different meiotic cells (Tsai et al, 2011). Time-lapse analyses in living spermatocytes using GFP-tagged chromosomal sites could be useful for revealing such stable connections; a stable connection site that happened to lie sufficiently close to a tagged chromosomal site should restrict the relative mobility of the tagged homologous alleles, perhaps dramatically so in favorable cases.…”

“…Using a GFP-tagged centromere protein to visualize centromeres in live cells, centromeres have been found to cluster non-specifically in early prophase, when euchromatic sequences are tightly paired, then to sort into pairwise and strictly homologous associations shortly after the loss of homologous pairing in the chromosome arms (Vazquez et al, 2002;Yan et al, 2010). A recent FISH study demonstrated that this pairing is centromere-specific and does not extend even into very nearby pericentromeric heterochromatin (heterochromatin situated near to a centromere) (Tsai et al, 2011), and thus could be an example of PC-like behavior. However, homologous centromere pairing is short-lived; centromeres become unpaired by mid-prophase I and remain unpaired throughout the remainder of meiosis I.…”

Nucleolar activity and CENP-C regulate CENP-A and CAL1 availability for centromere assembly in meiosis

“…Given that rDNA loci are located on X and Y chromosomes (McKee et al, 2012), we hypothesized that X and Y centromeres associate with nucleoli and might aberrantly assemble CENP-A. Using fluorescent in situ hybridisation (FISH) probes specific for repeat sequences on the X (359 bp satellite) or Y (AATAC satellite) chromosomes (Tsai et al, 2011), and coimmunostaining for Fibrillarin, we confirmed that X and Y chromosomes associate with nucleoli in wild-type control and Cenp-C Z3-4375 /TM3 spermatocytes (Fig. 4A).…”

“…However, our use of FISH probes to identify X and Y chromosomes, combined with antibody staining of endogenous CENP-A, could explain discrepancies between the two studies. It is also possible that the high CENP-A observed on sex chromosomes is sometimes due to their close association with the small fourth chromosome (Tsai et al, 2011), which is also located proximal to nucleoli. Interestingly, we find that in Cenp-C mutants the highly elevated CENP-A at X or Y centromeres is not retained past prophase I, i.e.…”