Skip to Content
Merck
  • Regulation of PKD1-mediated Golgi to cell surface trafficking by Gαq subunits.

Regulation of PKD1-mediated Golgi to cell surface trafficking by Gαq subunits.

Biology of the cell (2013-11-02)
A Soledad Coria, M Luján Masseroni, Alberto M Díaz Añel
ABSTRACT

Heterotrimeric GTP-binding proteins play a key role in cell trafficking regulation. Above all, specific Gβγ subunits have been shown to be a major component of a signal transduction pathway, which also involves phospholipases C (PLC), protein kinases C (PKC) and D (PKD), whose main function is to regulate transport between Golgi and plasma membrane. It was the involvement of PLC which led us to study the role of the other member of this G protein family, the α subunits, in the regulation of membrane fission at the Golgi apparatus. Among constitutive active (QL) variants of different G protein α subunit sub-families, only GαqQL subunits were able to induce Golgi fragmentation, a phenotype that mainly reflects a membrane fission increase at this organelle. This phenotype was not observed with a GαqQL palmitoylation mutant, showing the need for a membrane-bounded subunit. Besides, GαqQL-dependent Golgi fission was blocked by specific PLC and PKC inhibitors, and in the presence of a PKD1-kinase dead variant. In addition, GαqQL was the only α subunit capable of inducing PKD1 phosphorylation. Finally, Vesicular Stomatitis Virus thermosensitive mutant glycoprotein (VSVG tsO45) transport assays have demonstrated that GαqQL acts directly on Golgi membranes to regulate trafficking between this organelle and plasma membrane. All these results indicate Gαq subunits for the first time as a regulator of PKD-mediated intracellular trafficking between Golgi apparatus and plasma membrane, opening new perspectives in the understanding of internal trafficking regulation by external signals through G protein-coupled receptors.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
U-73122, U-73122, CAS 112648-68-7, inhibits agonist-induced phospholipase C activation (IC50 = 1-2.1 µM) in human platelets and neutrophils.
Sigma-Aldrich
Bisindolylmaleimide I, A highly selective, cell-permeable, and reversible protein kinase C (PKC) inhibitor (IC₅₀ = 10 nM) that is structurally similar to staurosporine.
Sigma-Aldrich
Monoclonal Anti-Tubulin, Tyrosine antibody produced in mouse, clone TUB-1A2, ascites fluid
Sigma-Aldrich
Anti-TGN46 antibody produced in rabbit, ~1 mg/mL, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
Cycloheximide solution, Ready-Made Solution, microbial, 100 mg/mL in DMSO, Suitable for cell culture
Sigma-Aldrich
Monoclonal Anti-VSV Glycoprotein−Cy3 antibody produced in mouse, clone P5D4, purified immunoglobulin, buffered aqueous solution