Identification of Ce-AF-6, a novel Caenorhabditis elegans protein, as a putative Ras effector

Mammalian Ras proteins associate with multiple effecters, including Raf, Ra l guanine nucleotide dissociation stimulator, phosphoinositide 3-kinase and AF-6. In the nematode Caenorhabditis elegans, LIN-45/Raf has been identifi ed genetically as an effector of LET-60/Ras. To search for other effecters in C. elegans, we carried out a yeast two-hybrid screening for LET-60-assoc iating proteins. The screening identified a novel protein, designated Ce-AF -6, which exhibited a strong structural homology with human AF-6, rat Afadi n and Drosophila melanogaster Canoe and possessed both the Ras-associating (RA) domain and the PSD-95/D1gA/ZO-1 (PDZ) domain. Ce-AF-6 associated with human Ha-Ras in a GTP-dependent manner, with an efficiency comparable to th at of human Raf-1 Ras-binding domain. When the effects of mutations of the Ras effector region residues were examined for associations with various ef fecters, Ce-AF-6 was found to possess a distinct and the most rigorous requ irement for the effector region residues. These results strongly suggest th at Ce-AF-6 is a putative effector of Ras that possesses a distinct recognit ion mechanism for association with Ras. (C) 1998 Elsevier Science B.V. All rights reserved.