MAMMALIAN DNA-LIGASE-III - MOLECULAR-CLONING, CHROMOSOMAL LOCALIZATION, AND EXPRESSION IN SPERMATOCYTES UNDERGOING MEIOTIC RECOMBINATION

Three biochemically distinct DNA ligase activities have been identifie d in mammalian cell extracts, We have recently purified DNA ligase II and DNA ligase III to near homogeneity from bovine liver and testis ti ssue, respectively, Amino acid sequencing studies indicated that these enzymes are encoded by the same gene, In the present study, human and murine cDNA clones encoding DNA ligase III were isolated with probes based on the peptide sequences, The human DNA ligase III cDNA encodes a polypeptide of 862 amino acids, whose sequence is more closely relat ed to those of the DNA ligases encoded by poxviruses than to replicati ve DNA ligases, such as human DNA ligase I, In vitro transcription and translation of the cDNA produced a catalytically active DNA ligase si milar in size and substrate specificity to the purified bovine enzyme. The DNA ligase III gene was localized to human chromosome 17, which e liminated this gene as a candidate for the cancer-prone disease Bloom syndrome that is associated with DNA joining abnormalities, DNA ligase III is ubiquitously expressed at low levels, except in the testes, in which the steady-state levels of DNA ligase III mRNA are at least 10- fold higher than those detected in other tissues and cells, Since DNA ligase I mRNA is also present at high levels in the testes, we examine d the expression of the DNA ligase genes during spermatogenesis, DNA l igase I mRNA expression correlated with the contribution of proliferat ing spermatogonia cells to the testes, in agreement with the previousl y defined role of this enzyme in DNA replication, In contrast, elevate d levels of DNA ligase III mRNA were observed in primary spermatocytes undergoing recombination prior to the first meiotic division. Therefo re, we suggest that DNA ligase III seals DNA strand breaks that arise during the process of meiotic recombination in germ cells and as a con sequence of DNA damage in somatic cells.