Mutational analysis of phospholipase C‐β2

Open Access
Publisher Website
Google Scholar
Abstract
Members of the β isozyme subfamily of the phosphoinositide‐specific phospholipases C (PLCβ) have recently been shown to be stimulated by both guanine‐nucleotide‐binding protein α and βγ subunits. The α subunits of the Gq class activate PLCβ isozymes in the order of PLCβ1≥ PLCβ3≫ PLCβ2, which is different from the order of PLCβ3 > PLCβ2 > PLCβ1 for βγ subunit stimulation. The C‐terminal region of PLCβ1, in particular the sequence between Thr903 and Leu1142, has been shown to be involved in interacting with activated αq subunits and to contain a region required for efficient membrane association of PLCβ1 [Park, D., Jhon, D.‐Y., Lee, C.‐W., Ryu, S. H. & Rhee, S. G. (1993) J. Biol. Chem. 268, 3710–3714, and Wu, D., Jiang, H., Katz, A. & Simon, M. I. (1993) J. Biol. Chem. 268, 3704–3709]. To examine the structure‐function relationships of a PLCβ isozyme highly sensitive to βγ subunit stimulation, we have altered the cDNA of PLCβ2 by site‐directed mutagenesis and have examined the effects of these structural alterations on the functional properties of the mutant polypeptides. The results show that the C‐terminal region of PLCβ2 downstream of Phe818, which corresponds to Tyr816 of PLCβ1, contains a region essential for membrane association, but is required neither for the interaction of PLCβ2 with Ca2+ and the phospholipid substrate, nor for βγ subunit stimulation of PLCβ2. These data suggest that PLCβ isozymes are activated by αq and βγ subunits via distinct domains.