702B Poster - 11. Cell division and cell growth
Friday April 08, 2:00 PM - 4:00 PM

Authors:
Susan McMahan; Colette Anikwue; Jeff Sekelsky

Affiliation: University of North Carolina at Chapel Hill

Keywords:
b. meiosis; c. meiosis

Sturtevant reported in his classic 1913 paper¹ that the presence of one meiotic crossover (CO) discourages others, a phenomenon Muller termed interference. More than 100 years later we still don’t know the mechanism of interference. We propose a model in which induced phase separation of pro-CO proteins from the synaptonemal complex (SC), a protein structure that forms between paired homologous chromosomes, followed by coarsening, these pro-CO proteins to accumulate at a single site or widely separated sites.

In Drosophila, ~20-24 double-strand breaks (DSBs) are made per meiosis, distributed uniformly throughout the euchromatin (at large scale). About 1/4 of these are designated to become COs and the rest are repaired as non-crossovers. In her EM studies of recombination Drosophila, Carpenter² discovered a CO-associated structure she called the recombination nodule (RN), a 100 nM sphere that sits on top of the SC. The proposal by Dernburg and colleagues that the SC had liquid-like properties³ and that recombination factors can move within this compartment⁴ suggest the RN may be a droplet containing CO-designating proteins.

We propose that the pro-CO mei-MCM complex (Mei-217, Mei-218, and Rec)⁵ is localized to DSB sites. A 2^nd set of pro-CO factors, the RING-finger proteins Vilya, Narya, and Nenya^6,7 (here abbreviated VNN), move within the liquid SC. Phosphorylation of the intrinsically disordered domain of Mei-218 by Mei-41 (ATR kinase) promotes physical interaction with VNN. VNN accumulates and dewets from the SC. Exchange between nearby occurs through the SC, but larger accumulations become more stable, so that each SC (i.e., each chromosome arm) eventually has a single VNN or widely separated VNN foci. These sites are designated as COs. This process naturally leads to crossover assurance - the guarantee that each pair of homologous chromosomes (excepting the 4 in Drosophila melanogaster) almost always get at least one crossover to promote accurate segregation. In our view, interference is merely a consequence of the assurance process.

We made numerous in situ mutations in mei-218 to alter ATR phosphorylation sites. Mutation of S/T to the phosphomimetic residue D results in decreased COs and loss of interference, with increasing severity for 3, 5, or 8 sites mutated. In the most severe case there is high negative interference, where double-COs in two adjacent intervals are significantly more frequent than expected if the intervals are independent of one another. In addition to these data, progress toward dissection of different phosphorylation sites and mapping of interactions between VNN proteins and phospho-Mei-218 will be presented.

¹ Sturtevant 1913 J Exp Biol.; ² Carpenter 1979 Chromosoma; ³ Rog et al. 2017 Elife; ⁴ Zhang et al. 2018 Elife. ⁵ Kohl et al. 2012 Science; ⁶ Lake et al. 2015 eLife; ⁷ Lake et al. 2019 PLOS Gen