A NOVEL FAMILY OF SERINE THREONINE KINASES PARTICIPATING IN SPERMIOGENESIS/

The molecular mechanisms regulating the spectacular cytodifferentiatio n observed during spermiogenesis are poorly understood. We have recent ly identified a murine testis-specific serine kinase (tssk) 1, constit uting a novel subfamily of serine/threonine kinases. Using low stringe ncy screening we have isolated and molecularly characterized a second closely related family member, tssk 2, which is probably the orthologu e of the human DGS-G gene. Expression of tssk 1 and tssk 2 was limited to the testis of sexually mature males. Immunohistochemical staining localized both kinases to the cytoplasm of late spermatids and to stru ctures resembling residual bodies. tssk 1 and tssk 2 were absent in re leased sperms in the lumen of the seminiferous tubules and the epididy mis, demonstrating a tight window of expression restricted to the last stages of spermatid maturation. In vitro kinase assays of immunopreci pitates containing either tssk 1 or tssk 2 revealed no auto-phosphoryl ation of the kinases, however, they led to serine phosphorylation of a coprecipitating protein of similar to 65 kD. A search for interacting proteins using the yeast two-hybrid system with tssk 1 and tssk 2 cDN A as baits and a prey cDNA library from mouse testis, led to the isola tion of novel cDNA, interacting specifically with both tssk 1 and tssk 2, and encoding the coprecipitated 65-kD protein phosphorylated by bo th kinases. Interestingly, expression of the interacting clone was als o testis specific and paralleled the developmental expression observed for the kinases themselves. These results represent the first demonst ration of the involvement of a distinct kinase family, the tssk serine /threonine kinases, together with a substrate in the cytodifferentiati on of late spermatids to sperms.