Phospholipase Cδ1 Is a Guanine Nucleotide Exchanging Factor for Transglutaminase II (Gαh) and Promotes α1B-Adrenoreceptor-mediated GTP Binding and Intracellular Calcium Release

Abstract

Effectors involved in G protein-coupled receptor signaling modulate activity of GTPases through GTPaseactivating protein or guanine nucleotide exchanging factor (GEF). Phospholipase C␦1 (PLC␦1) is an effector in tissue transglutaminase (TGII)-mediated ␣ 1B -adrenoreceptor (␣ 1B AR) signaling. We investigated whether PLC␦1 modulates TGII activity. PLC␦1 stimulated GDP release from TGII in a concentration-dependent manner, resulting in an increase in GTP␥S binding to TGII. PLC␦1 also inhibited GTP hydrolysis by TGII that was independent from the ␣ 1B AR. These results indicate that PLC␦1 is GEF for TGII and stabilizes the GTP⅐TGII complex. When GEF function of PLC␦1 was compared with that of the ␣ 1B AR, the ␣ 1B AR-mediated GTP␥S binding to TGII was greater than PLC␦1-mediated binding and was accelerated in the presence of PLC␦1. Thus, the ␣ 1B AR is the prime GEF for TGII, and GEF activity of PLC␦1 promotes coupling efficacy of this signaling system. Overexpression of TGII and its mutants with and without PLC␦1 resulted in an increase in ␣ 1B AR-stimulated Ca 2؉ release from intracellular stores in a TGIIspecific manner. We conclude that PLC␦1 assists the ␣ 1B AR function through its GEF action and is primarily activated by the coupling of TGII to the cognate receptors.

Direct Activation of Phospholipase C-ϵ by Rho

“…1). As observed previously, transfection of PLC-⑀ alone resulted in a small increase in [ 3 H]inositol phosphates that was substantially augmented by cotransfection with G␤ 1 ␥ 2 or GTPase-deficient mutants of G␣ 12 , G␣ 13 , or H-Ras (Fig 1A). Surprisingly, constitutively active forms of RhoA, RhoB, and RhoC also markedly stimulated inositol phosphate accumulation after cotransfection with PLC-⑀ ( Fig 1B, and data not shown).…”

“…Identification of an Insertion in the Y Box of PLC-⑀ -As illustrated in the experiments above, PLC-⑀ is unique among PLC isozymes in requiring only the catalytic core for activation by Rho, G␣ 12 , and G␣ 13 . To identify unique regions within the catalytic core of PLC-⑀ that potentially could impart this responsiveness, pair-wise and multiple sequence alignments were performed comparing PLC-⑀ isoforms with the other …”

“…The heterotrimeric G protein subunits G␣ q (3)(4)(5)(6) and G␤␥ (7-9) directly activate PLC-␤ isozymes, whereas the activity of PLC-␥ isozymes is increased via phosphorylation and translocation as a consequence of the activation of tyrosine kinase receptors (10 -12). The mechanism(s) of regulation of PLC-␦ is less clear, but it has been suggested to involve Gh proteins (13) as well as Ca 2ϩ mobilization (14). A fourth member of the family of PLC enzymes, PLC-⑀, was identified recently (15)(16)(17)(18).…”

Tissue transglutaminase expression promotes cell attachment, invasion and survival in breast cancer cells

“…In addition to catalysing the calcium-dependent post-translation modification of proteins, TG2 can also catalyse calciumindependent hydrolysis of guanosine triphosphate (GTP) and adenosine triphosphate, the protein disulfide isomerase reaction (Chandrashekar et al, 1998) and serine/threonine kinase activity (Chen and Mehta, 1999;Fesus and Piacentini, 2002;Lorand and Graham, 2003;Mishra and Murphy, 2004). The ability of TG2 to hydrolyse GTP enables it to serve as a signaling molecule in transmitting outside signal from a 1b adrenergic receptors to a downstream cytoplasmic target, phospholipase C (Baek et al, 2001). Although, predominantly a cytosolic protein, TG2 can also be secreted outside the cell where it regulates cell-matrix interactions (Aeschlimann and Thomazy, 2000); can translocate to the nucleus where it associates with pRb, p53 and histones to regulate certain cellular functions (Milakovic et al, 2004;; and can be expressed on the cell membrane in association with b-integrins where it serves as a coreceptor for integrin-mediated binding to fibronectin (Fn) (Akimov and Belkin, 2001).…”

Activation of Phospholipase C-ε by Heterotrimeric G Protein βγ-Subunits

“…PLC-␥ isozymes are activated by protein phosphorylation following from activation of tyrosine kinase receptors or from receptors linked to tyrosine kinases (10 -12). Although there is evidence for Ca 2ϩ -and transglutaminase II-promoted regulation of PLC-␦ (13,14), the importance of hormonal regulation of this PLC isozyme remains unclear (15).…”