Protein 4.1 R-135 interacts with a novel centrosomal protein (CPAP) which is associated with the gamma-tubulin complex

Using a yeast two-hybrid system, we isolated a novel human centrosomal prot ein, CPAP (centrosomal P4.1-associated protein), which specifically interac ts with the head domain of the 135-kDa protein 4.1R isoform (4.1R-135). Seq uence analysis revealed that the carboxyl terminus of CPAP has 31.3% amino acid identity with human Tcp-10 (a t-complex responder gene product). Inter estingly, most of the sequence identity is restricted to two conserved regi ons. One carries a leucine zipper, which may form a series of heptad repeat s involved in coiled-coil formation; the other contains unusual glycine rep eats with unknown function. Immunofluorescence analysis revealed that CPAP and gamma-tubulin are localized within the centrosome throughout the cell c ycle. CPAP cosediments with gamma-tubulin in sucrose gradients and coimmuno precipitates with gamma-tubulin, indicating that CPAP is a part of the gamm a-tubulin complex. Furthermore, functional analysis revealed that CPAP is l ocalized within the center of microtubule asters and may participate in mic rotubule nucleation. The formation of microtubule asters was significantly inhibited by anti-CPAP antibody. Together, these observations indicate that CPAP may play an important role in cell division and centrosome function.