DNA replication may be uncoupled from nuclear and cellular division in temperature-sensitive embryonic lethal mutants of Caenorhabditis elegans

Abstract

The nuclear and DNA contents of 18 temperature-sensitive embryonic lethal mutants of Caenorhabditis elegans have been determined at the time of arrest. After each mutant was shifted to restrictive temperature, the embryonic arrest stage was recorded by the number of nuclei counted in embryonic squashes and as DNA nuclear equivalents recorded by flow cytometry. Together the two methods complemented each other and provided qualitative and quantitative information concerning nuclear number, DNA content, morphological stage of arrest, and presence of anucleate embryonic cells. The arrest stage for most of the embryonic lethal mutants demonstrated their potential to continue nuclear division and DNA replication beyond their respective temperature-sensitive periods. These results suggested that the mutants' primary defect did not reside on a pathway closely coupled to DNA replication or nuclear division. For example, B65 contained up to 650 DNA nuclear equivalents beyond the end of its temperature-sensitive period at the 20- to 30-cell stage. B65 is of additional interest because it continued DNA synthesis beyond the normal 550-cell stage and at the same time failed to progress beyond the morphogenetic period. The presence of enlarged nuclei that contained extra DNA demonstrated that DNA replication was independent of nuclear and cellular division. For example, B1 and B244 arrested with 1 to 50 and 1 to 200 nuclei, respectively. However, they both contained 250 to 400 nuclear equivalents of DNA. Conversely, the presence of embryonic cells without nuclei suggested that cell division may also be independent of nuclear division or nuclear migration. The enlarged nuclear and anucleate embryonic cells were observed only in those mutants requiring a normal parental function.