hCASK and hDlg associate in epithelia, and their Src homology 3 and guanylate kinase domains participate in both intramolecular and intermolecular interactions

Membrane associated guanylate kinase (MAGUK) proteins act as molecular scaf folds organizing multiprotein complexes at specialized regions of the plasm a membrane. All MAGUKs contain a Src homology 3 (SH3) domain and a region h omologous to yeast guanylate kinase (GUK). We showed previously that one MA GUK protein, human CASK (hCASK), is widely expressed and associated with ep ithelial basolateral plasma membranes. We now report that hCASK binds anoth er MAGUK, human discs large (hDlg). Immunofluorescence microscopy demonstra tes that hCASK and hDlg colocalize at basolateral membranes of epithelial c ells in small and large intestine. These proteins co-precipitate from lysat es of an intestinal cell line, Caco-2, The GUK domain of hCASK binds the SH 3 domain of hDlg in both yeast two-hybrid and fusion protein binding assays , and it is required for interaction with hDlg in transfected HEK293 cells. In addition, the SH3 and GUK domains of each protein participate in intram olecular binding that in vitro predominates over intermolecular binding. Th e SH3 and GUK domains of human p55 display the same interactions in yeast t wo-hybrid assays as those of hCASK. Not all SH3-GUK interactions among thes e MAGUKs are permissible, however, implying specificity to SH3-GUK interact ions in vivo. These results suggest MAGUK scaffold assembly may be regulate d through effects on intramolecular SH3-GUK binding.