G2 regulatory processes require speedbump for Wee1 kinase activity

Abstract

Wee1 is a conserved Cdk1 inhibitory kinase operating at the G2/M checkpoint to prevent entry into mitosis until the genome has been surveilled and replication is complete. We report here that the early arrest mutant speedbump is a loss-of-function mutation in the zebrafish ortholog of wee1. Like other creatures lacking Wee1 kinase, cells in the mutant enter mitosis early. Eventually, mutant cells exhibit chromosomal defects and undergo apoptosis. Live recordings of the mutant reveal that as gastrula cells transition from maternal to zygotic control, their cell cycle gets progressively shorter rather than lengthening as seen in wild-type embryos. This suggests that Wee1 kinase inhibition is part of a mechanism to slow the cell cycle that we posit is independent of its role in blocking entry into mitosis to prevent DNA damage. Supporting this view, we show that Wee1 kinase is also crucial for tissues that normally exit the cell cycle in the G2 phase. In the absence of Wee1 kinase, hatching gland cells, which typically cease dividing before speedbump defects appear, no longer remain in G2, and instead advance into mitosis before prematurely dying. Finally, we demonstrate that Wee1 kinase is essential for the endoreplication cycle in the yolk cell. We show that wild-type yolk cell nuclei transition to an S and G endocycle after they cease mitosis in the blastula. However, without Wee1 kinase these nuclei have difficulty attaining this endocycle and sometimes regress back into mitosis. We conclude that besides the regulation of mitotic timing, Wee1 kinase has other G2 regulatory roles not previously reported in which controlling entry into mitosis must be coordinated with other cellular processes.