Replication protein A (AtRPA1a) is required for class I crossover formation but is dispensable for meiotic DNA break repair.

Replication protein A (RPA) is involved in many aspects of DNA metabolism including meiotic recombination. Many species possess a single RPA1 gene but Arabidopsis possesses five RPA1 paralogues. This feature has enabled us to gain further insight into the meiotic role of RPA1. Proteomic analysis implicated one of the AtRPA1 family (AtRPA1a) in meiosis. Immunofluorescence studies confirmed that AtRPA1a is associated with meiotic chromosomes from leptotene through to early pachytene. Analysis of an Atrpa1a mutant revealed that AtRPA1a is not essential at early stages in the recombination pathway. DNA double-strand breaks are repaired in Atrpa1a, but the mutant is defective in the formation of crossovers, exhibiting a 60% reduction in chiasma frequency. Consistent with this, localization of recombination proteins AtRAD51 and AtMSH4 appears normal, whereas the numbers of AtMLH1 and AtMLH3 foci at pachytene are significantly reduced. This suggests that the defect in Atrpa1a is manifested at the stage of second-end capture. Analysis of Atrpa1a/Atmsh4 and Atrpa1a/Atmlh3 double mutants indicates that loss of AtRPA1a predominantly affects the formation of class I, interference-dependent crossovers.