IDENTIFICATION OF HUMAN LIVER CDNA CLONES WHOSE PRODUCTS INTERACT WITH G-PROTEIN BETA-SUBUNIT OF SCHIZOSACCHAROMYCES-POMBE

Heterotrimeric GTP binding proteins (G-proteins) are composed of alpha , beta, and gamma subunits and have important roles in the cell signal ling from cell surface receptors to effecters. The alpha subunit as we ll as the beta gamma complex are known to interact with effector molec ules to deliver specific signals to downstream genes. In an attempt to identify the molecules interacting directly with G-protein beta subun it (G beta) in signal pathway, G-protein beta subunit gene of Schizosa ccharomyces pombe was used as bait to screen a human liver cDNA librar y in a yeast two-hybrid system. When the coding region DNA of the S. p ombe beta subunit gene, gpb1(+), was cloned into the 3'-end of the GAL 4 DNA binding domain sequence and used to screen a human liver cDNA li brary cloned at the 3'-end of the GAL4 activation domain, 42 novel clo nes showing beta-galactosidase activity were obtained from the 9x10(6) transformants. The sequence analysis of these clones revealed that th e four clones contain sequence homologies with a receptor tyrosine kin ase, a mammalian homolog of hsp40 (DNAJ), a NMDA receptor glutamate-bi nding protein, and a protein serine kinase. These four clones showed s pecific binding to G-protein beta subunit of S. pombe in vitro. The ch aracterization of these proteins in the G-protein-mediated signal path way will be useful in elucidating the precise mechanism of G-protein f unction.