Genetic probing of homologous recombination and non-homologous end joining during meiotic prophase in irradiated mouse spermatocytes

This study was designed to obtain a better insight into the relative contribution of homologous recombination (HR) and non-homologous end joining (NHEJ) to the repair of radiation-Induced DNA double-strand breaks (DSBs) at first meiotic prophase. Early and late pachytene and early diplotene spermatocytes that had completed crossing over were sampled. We studied the kinetics of gamma-H2AX chromatin foci removal after irradiation of mice deficient for HR and mice deficient for NHEJ Analyzing gamma-H2AX signals in unirradiated RAD54/RAD54B deficient spermatocytes indicated Incomplete meiotic recombination repair due to the pronounced increase of gamma-H2AX foci in late prophase primary spermatocytes In these mice, 8 h after irradiation, early pachytene spermatocytes showed a reduction of the numbers of gamma-H2AX foci by 52% compared to 82% in the wild type, the difference being significant. However, after crossing over (In late pachytene and early diplotene), no effect of RAD54/RAD54B deficiency on the reduction of irradiation-induced foci was observed In NHEJ deficient SCID mice, repair kinetics in early spermatocytes were similar to those in wild type mice However, 1 h after irradiation in late pachytene and early diplotene spermatocytes 1.7 times more foci were found than in wild type mice. This difference might be related to the absence of a DNA-PKcs dependent fast repair component in SCID mice. As subsequent repair is normal, HR likely is taking over Taken together, the results obtained in RAD54/RAD54B deficient mice and in SCID mice indicate that DSB repair in early pachytene spermatocytes is mainly carried out through HR In late spermatocytes (late pachytenes and early diplotenes) NHEJ is active However, probably there is an interplay between these repair pathways and when in late spermatocytes the NHEJ pathway is compromised HR may take over (C) 2010 Elsevier B V All rights reserved.