Optogenetic control of PLC-γ1 activity polarizes cell motility

Area of Science:

• Cell Biology
• Biochemistry
• Cancer Research

Background:

• Phospholipase C-γ1 (PLC-γ1) signaling is crucial for mesenchymal cell movement.
• Understanding PLC-γ1's sufficiency in directing cell motility requires dissecting its autoinhibition and activity.
• Previous studies faced challenges in isolating PLC-γ1's specific contribution due to basal autoinhibition.

Purpose of the Study:

• To investigate whether PLC-γ1 signaling is sufficient to bias mesenchymal cell motility.
• To explore the regulatory logic of PLC-γ1 using cancer-associated mutations.
• To determine if local membrane recruitment of PLC-γ1 can activate substrate hydrolysis and cell motility.

Main Methods:

• Utilized optogenetic control (OptoPLC-γ1) for light-induced membrane recruitment of PLC-γ1 in Plcg1-null fibroblasts.
• Employed cancer-associated mutations (P867R, S345F, D1165H) to study PLC-γ1 regulation and activity.
• Assessed cell motility responses, phosphorylation status (Tyr783), and lipase activity upon OptoPLC-γ1 recruitment.

Main Results:

• Basal autoinhibition of PLC-γ1 complicates assessment of its activity.
• Phosphorylated Tyr783 (pTyr783) is a marker of dysregulated autoinhibition, not enzyme activity level.
• Deregulated OptoPLC-γ1 mutants (P867R, S345F, D1165H) showed increased phosphorylation and, upon membrane localization, activated hydrolysis and motility.
• Local recruitment of OptoPLC-γ1 S345F specifically polarized cell motility on demand, demonstrating spatial dose-sensitivity.
• This lipase-dependent motility response was only partially inhibited by blocking canonical PLC-γ1 pathways.

Conclusions:

• The canonical hallmark of PLC-γ1 activity (pTyr783) reflects dysregulated autoinhibition, not activity level.
• Local membrane recruitment of deregulated PLC-γ1 mutants is sufficient to drive substrate hydrolysis and cell motility.
• Local activation of PLC-γ1 is sufficient to direct cell motility, reframing the understanding of its regulation.