IDENTIFICATION OF NOVEL HUMAN WW DOMAIN-CONTAINING PROTEINS BY CLONING OF LIGAND TARGETS

A recently described protein module consisting of 35-40 semiconserved residues, termed the WW domain, has been identified in a number of div erse proteins in eluding dystrophin and Yes-associated protein (YAP). Two putative ligands of YAP, termed WBP-1 and WBP-2, have been found p reviously to contain several short peptide regions consisting of PPPPY residues (PY motif) that mediate binding to the WW domain of YAP. Alt hough the function(s) of the WW domain remain to be elucidated, these observations strongly support a role for the WW domain in protein-prot ein interactions. Here we report the isolation of three novel human cD NAs encoding a total of nine WW domains, using a newly developed appro ach termed COLT (cloning of ligand targets), in which the rapid clonin g of modular protein domains is accomplished by screening cDNA express ion libraries with specific peptide ligands. Two of the new genes iden tified appear to be members of a family of proteins, including Rsp5 an d Nedd-4, which have ubiquitin-protein ligase activity. In addition, w e demonstrate that peptides corresponding to PY and PY-like motifs pre sent in several known signaling or regulatory proteins, including RasG AP, AP-2, p53BP-2 (p53-binding protein-2), interleukin-6 receptor-alph a, chloride channel CLCN5, and epithelial sodium channel ENaC, can sel ectively bind to certain of these novel WW domains.