Role of amphibian egg transglutaminase in the development of secondary cytostatic factor in vitro.
Fresh cytosols extracted from unfertilized amphibian eggs contain a cytostatic factor (CSF) which arrests the cell cycle at metaphase when microinjected into cleaving blastomeres. This CSF is sensitive to Ca2+, and is designated primary CSF (1 degree CSF). During storage of Ca(2+)-containing cytosols at 2 degrees C, stable CSF activity appears, designated secondary CSF (2 degrees CSF). In Rana pipiens egg cytosols, the development of 2 degrees CSF coincides with the formation of a protein complex with a molecular weight above 2,000 kDa, and this large molecule exhibits a high 2 degrees CSF activity when purified (Shibuya and Masui, 1989: Development 106:799-808). The present study shows that both the formation of 2 degrees CSF protein complex and the development of its activity are inhibited by ethylamine and glycine-ethyl-ester (GEE), both known as potent transglutaminase (TGase) inhibitors. An affinity-purified polyclonal antibody raised against mammalian transglutaminase reacts with an approximately 68-kDa protein in fresh egg cytosols, as well as with the 2 degrees CSF protein complex. In cytosols deprived of transglutaminase by immunoprecipitation, neither the development of 2 degrees CSF activity nor the formation of its protein complex can occur. These results indicate that transglutaminase of Rana pipiens eggs is responsible or the formation of 2 degrees CSF, and that transglutaminase itself is incorporated into 2 degrees CSF molecules.
Zhang
J
J
Department of Zoology, University of Toronto, Ontario, Canada.
Masui
Y
Y
D000818:Animals; D002118:Calcium / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005021:Ethylamines / Q000494:pharmacology; D005998:Glycine / Q000031:analogs & derivatives / Q000494:pharmacology; D066298:In Vitro Techniques; D008677:Metaphase; D009865:Oocytes / Q000378:metabolism*; D017231:Proto-Oncogene Proteins c-mos / Q000378:metabolism*; D011894:Rana pipiens; D011503:Transglutaminases / Q000378:metabolism*
D004791:Enzyme Inhibitors; D005021:Ethylamines; C022010:glycine ethyl ester; D011503:Transglutaminases; D017231:Proto-Oncogene Proteins c-mos; D002118:Calcium; D005998:Glycine; C041564:ethylamine