Observation of Extensive Chromosome Axis Remodeling during the “Diffuse-Phase” of Meiosis in Large Genome Cereals

Abstract

The production of balanced fertile haploid gametes requires the faithful separation of paired (synapsed) chromosomes toward the end of meiotic prophase I (desynapsis). This involves the timely dissolution of the synaptonemal complex during the pachytene-diplotene transition, a stage traditionally referred to as the “diffuse stage.” In species with large genomes such as, barley (Hordeum vulgare L.) and wheat (Triticum aestivum L.) we know most about the early stages of meiotic prophase I. There, synapsis initiates at the telomeric ends of chromosomes and progresses toward the centromeric regions through the ordered assembly of the synaptonemal complex (SC). Synapsis is impacted by recombination (crossing over, CO) which locally modifies the extent of chromatin compaction and extension. CO is uneven along the chromosomes, occurring mainly toward the telomeric regions resulting in a highly skewed distribution of recombination events. However, we know very little about the process of desynapsis which occurs during the “diffuse stage,” where the synapsed and recombined chromosomes faithfully desynapse and separate into daughter cells. Here, using 3D-SIM super-resolution immuno-cytology combined with the use of antibodies directed against two crucial SC proteins, ASY1 and ZYP1, we followed the whole of meiosis I (i.e., both synapsis and desynapsis) in both barley and wheat. We showed that synapsis forms a characteristic tri-partite SC structure in zygotene (more clearly seen in barley). Toward the end of meiosis I, as the SC starts to disassemble, we show that extensive chromosome axis remodeling results in the formation of characteristic “tinsel-like” structures in both wheat and barley. By using a mutant (des10) that is severely compromised in polymerization of ZYP1during synapsis, we show that tinsel structure formation during SC dissolution is not dependant on full synapsis and may relate instead to changes in expansion stress. Our observations highlight a potentially new role for ASYNAPSIS1 (ASY1) in desynapsis, in addition to chromosome synapsis and cohesion.

Time-resolved transcriptome of barley anthers and meiocytes reveals robust and largely stable gene expression changes at meiosis entry

“…Pachytene and diplotene were somewhat easier to find based on anther size and acetocarmine spreads (Figure 1f) and were distinguished using antibodies against TaASY1 and HvZYP1 proteins (Figure 1g–h). Pachytene (Figure 1g) was characterized by the polymerization of HvZYP1 between the ASY1 axes along the entire chromosomes and diplotene (Figure 1h) was identifiable by the appearance of tinsel chromosomes as previously described (Colas et al , 2017). Since our primary focus was on synapsis and recombination, later stages from metaphase I to tetrads were combined into a single sample (Figure 1).…”

“…At leptotene, linear ASY1 axes are formed while there is no or little labelling of HvZYP1 at the initiation of synapsis (Figure 1d). Zygotene on the other hand was determined by the presence of HvZYP1 labelling during synapsis progression as previously described (Colas et al , 2017) (Figure 1e). Pachytene and diplotene were somewhat easier to find based on anther size and acetocarmine spreads (Figure 1f) and were distinguished using antibodies against TaASY1 and HvZYP1 proteins (Figure 1g–h).…”

“…Since our primary focus was on synapsis and recombination, later stages from metaphase I to tetrads were combined into a single sample (Figure 1). Metaphase I is characterised by the presence of seven ring bivalents (Figure 1i, j) while anaphase I (Figure 1k, l) and anaphase II samples show chromosomes or chromatin segregation, respectively (Colas et al , 2017). Anther samples of these later stages were not as synchronised as at prophase (leptotene–diplotene), containing both metaphase I and anaphase I or both anaphase II and telophase II.…”

“…Tubes were centrifuged at 4,500 rpm for 1 min and 20 μl meiocytes suspension was transferred onto a Polysine® 2 (Poly-L-Lysine coated slides) and left to air dry. The immuno-cytology was done according to Colas et al . (2017).…”

Meeting report – INDEPTH kick-off meeting

“…Offering a crop perspective, Isabelle Colas (James Hutton Institute, Dundee, UK) showed how 3D structured illumination microscopy (SIM) imaging has revealed novel features of chromatin organisation in meiotic chromosomes in barley that do not occur in Arabidopsis (Colas et al, 2017). She is using barley mutants and temperature variations to define the genetic and environmental factors that control both crossover frequency and location during meiosis, with the ultimate aim of providing improved insights for barley breeders.…”