Jf. Feng et al., alpha(1B)-adrenoceptor interacts with multiple sites of transglutaminase II: Characteristics of the interaction in binding and activation, BIOCHEM, 38(7), 1999, pp. 2224-2232
We previously reported that a novel GTP binding protein (G alpha(h)) is tis
sue type transglutaminase (TGII) and transmits the alpha(1B)-adrenoceptor (
AR) signal to phospholipase C (PLC) through its GTPase function. We have al
so shown that PLC-delta 1 is the effector in TGII-mediated signaling. In th
is study, interaction sires on TGII for the alpha(1B)-AR were identified us
ing a peptide approach and site-directed mutagenesis, including in vivo rec
onstitution of TGIIs with the alpha(1B)-AR and PLC-delta 1. To identify the
interaction sites, 11 synthetic peptides covering similar to 132 amino aci
d residues of the C-terminal domain of TGII were tested. The studies with t
he peptides revealed that three peptides, L-547-I-561, R-564-D-581, and Q(6
33)-E-646, disrupted formation of an alpha(1)-agonist-alpha(1B)-AR-TGII com
plex and blocked alpha(1B)-AR-mcdlated TGase inhibition in a dose-dependent
manner, indicating that these peptide regions are involved in recognition
and activation of TGII by the alpha(1B)-AR. These three regions were furthe
r evaluated with full-length TGIIs by constructing and coexpressing each si
te-directed mutant with the alpha(1B)-AR and PLC-delta 1 in COS-1 cells. Su
pporting the findings with these peptides, these TGII mutants lost 56-82% t
he receptor binding ability and reduced by 29-68% the level of alpha(1B)-AR
-mediated IP3 production via PLC-delta 1 as compared to those with wild-typ
e TGII. The results also revealed that the regions of R-564-D-581 and Q(633
)-E-646 were the high-affinity binding sites of TCII for the receptor and c
ritical for the activation of TGII by the receptor. Taken together, the stu
dies demonstrate that multiple regions of TGII interact with the alpha(1B)-
AR and that the alpha(1B)-AR stimulates PLC-delta 1 via TGII.