DIRECT INTERACTION BETWEEN YEAST SPINDLE POLE BODY COMPONENTS - KAR1PIS REQUIRED FOR CDC31P LOCALIZATION TO THE SPINDLE POLE BODY

The Saccharomyces cerevisiae genes KAR1 and CDC31 are required for the initial stages of spindle pole body (SPB) duplication in yeast. The C dc31 protein is most related to caltractin/centrin, a calcium-binding protein present in microtubule organizing centers in many organisms. B ecause of a variety of genetic interactions between CDC31 and KAR1 (Va llen, E. A., W. Ho, M. Winey, and M. D. Rose. 1994. Genetics. In press ), we wanted to determine whether Cdc31p and Kar1p physically interact . Cdc31p was expressed and purified from Escherichia coli and active f or binding calcium. Using a protein blotting technique, Cdc31p bound t o Kar1p in vitro via an essential domain in Kar1p required for SPB dup lication (Vallen, E. A., M. A. Hiller, T. Y. Scherson, and M. D. Rose. 1992a. J. Cell Biol. 117:1277-1287). By immunofluorescence microscopy , we determined that the interaction also occurs in vivo. Cdc31p was l ocalized to the SPB in wild-type cells but was mislocalized in a karl mutant strain. In a karl mutant containing a dominant CDC31 suppressor , Cdc31p was again localized to the SPB. Furthermore, the localization of Cdc31p to the SPB was affected by the overexpression of Kar1p-beta -galactosidase hybrids. Based on these data, we propose that the essen tial function of Kar1p is to localize Cdc31p to the SPB, and that this interaction is normally required for SPB duplication.