To identify proteins which interact with and potentially modulate the function of microtubules during spermatogenesis, we prepared a total testis MAP (microtubule-associated protein) antiserum and used it to isolate cDNA clones from a mouse testis cDNA expression library. Antibodies affinity purified by using one expression clone The process of cellular differentiation is intricately linked to morphological changes in the microtubular cytoskeleton. Microtubules, which form by self-assembly of a-and 3-tubulin heterodimers, require specific interactions with various proteins to establish diverse conformations and to perform their different functions. These interacting proteins are referred to as microtubule-associated proteins (MAPs). Two mechanisms can be envisaged for the involvement of MAPs in microtubule morphogenesis. First, the programmed expression of MAPs may be required for the terminal differentiation of a cell type; examples include MAP2 and tau in the determination of neuronal morphology (7). Second, the signal transduction pathway may influence microtubule organization via phosphorylation of tubulin and/or MAPs. Some kinases have been shown to colocalize with microtubules in vivo, providing a means for rapid phosphorylation of specific microtubule proteins. The c-mos product, which binds and phosphorylates tubulin (48), is an active component of the cytostatic factor, a factor isolated from mature Xenopus oocytes and thought to be responsible for the arrest of fertilized eggs at meiotic metaphase II (31). Among MAPs, phosphorylation of MAP2 has been extensively studied. Phosphorylation of MAP2 reduces its ability to stimulate tubulin assembly in vitro (20,35) and decreases its affinity for microtubules both in vitro (4) and in vivo (3). Type II cyclic AMP (cAMP)-dependent protein kinase phosphorylates and copurifies with brain MAP2 because of the high-affinity binding of RII to the amino terminus of the MAP2 molecule (38,44,45).Mammalian spermatogenesis is a continuum of cellular differentiation that has three principal phases: the mitotic proliferation and renewal of spermatogonia, the meiotic * Corresponding author. Electronic mail address: walden@ mcclbO.med.nyu.edu. divisions of spermatocytes, and spermiogenesis, a complex metamorphosis of the haploid germ cell culminating in the release of late spermatids into the lumen of the seminiferous tubule. During spermatogenesis the microtubular cytoskeleton exists in diverse conformations, including two microtubular arrays unique to spermiogenesis, namely the spermatid manchette and flagellum. The manchette temporally forms around the caudal nucleus of round spermatids and has been implicated in the subsequent process of sperm head shaping (9,32,39).We sought to identify proteins which interact with, and potentially modulate the function of, microtubules during spermatogenesis. We describe here the identification and characterization of one such protein, a novel 205-kDa microtubule-associated serine/threonine protein kinase, termed MAST205. MAST205 col...