Cell density sensing mediated by a G protein-coupled receptor activating phospholipase C

Area of Science:

• Cellular signaling in unicellular eukaryotes
• G protein-coupled receptor pathways
• Chemotaxis and cell-cell communication

Background:

• Dictyostelium discoideum uses conditioned medium factor (CMF) for density sensing during starvation.
• CMF signal transduction involves the chemoattractant cAMP receptor (cAR1) and Galpha2.
• The precise role of Galpha1 and phospholipase C (PLC) in CMF signaling was unclear.

Purpose of the Study:

• To elucidate the role of Galpha1 and PLC in conditioned medium factor (CMF) signaling.
• To understand how CMF regulates cAMP signal transduction.
• To investigate the mechanism of CMF-mediated cell aggregation.

Main Methods:

• Investigated CMF binding to membranes using guanosine 5'-3-O-(thio)triphosphate (GTPgammaS).
• Analyzed CMF signaling in genetically modified Dictyostelium discoideum cells lacking specific G protein subunits (Galpha1, Gbeta) or PLC.
• Measured PLC activity, IP3 levels, and cAMP-stimulated GTP hydrolysis rates.

Main Results:

• GTPgammaS inhibits CMF binding, suggesting a G protein-coupled receptor for CMF.
• Cells lacking Galpha1 do not show GTPgammaS inhibition of CMF binding or CMF regulation of cAMP signaling.
• CMF activates PLC, and this activation is dependent on Gbeta.
• Galpha1 null cells exhibit normal development and cAMP signaling in the presence of CMF.
• PLC-deficient cells show intermediate IP3 levels and GTP hydrolysis rates.

Conclusions:

• CMF signaling involves Galpha1 and Gbetagamma subunits, leading to PLC activation.
• Activated PLC inhibits Galpha2 GTPase, prolonging the cAMP-activated Galpha2-GTP state.
• This extended signaling cascade enables proper cAR1-mediated cAMP signal transduction and cell aggregation.