GTP-dependent interaction between phospholipase D and dynamin modulates fibronectin-induced cell spreading

Area of Science:

• Cell Biology
• Biochemistry
• Molecular Biology

Background:

• Phospholipase D (PLD) and dynamin are involved in cellular processes like endocytosis and actin cytoskeleton regulation.
• The precise mechanism linking PLD, dynamin, and fibronectin (FN)-induced cell spreading remains unclear.
• Understanding dynamin's regulatory role through protein interactions is an open question.

Purpose of the Study:

• To investigate the interplay between PLD and dynamin in fibronectin (FN)-mediated cell spreading.
• To determine if dynamin regulates PLD activity in a GTP-dependent manner.
• To elucidate the role of the PLD-dynamin interaction in cell signaling and cytoskeletal dynamics.

Main Methods:

• Utilized a GTP binding-deficient dynamin mutant (K44A) to assess GTP-dependent regulation.
• Inhibited PLD activity to evaluate its role in FN-induced cell spreading versus adhesion.
• Investigated PLD-dynamin interaction using a PLD mutant (R128K) lacking GAP activity.

Main Results:

• GTP-loaded dynamin mediates FN-induced cell spreading, while the mutant K44A does not.
• Inhibition of PLD activity blocks FN-induced cell spreading but not cell adhesion.
• PLD interacts with dynamin in a GTP-dependent manner during FN signaling, which is essential for PLD activation and cell spreading.
• A PLD mutant (R128K) lacking GAP activity enhances PLD-dynamin interaction, PLD activity, and cell spreading.

Conclusions:

• Dynamin acts as a GTP-dependent regulator of PLD activity.
• The interaction between GTP-loaded dynamin and PLD is critical for FN-induced cell spreading.
• Dynamin functions analogously to a small GTPase, mediating a "switch on" signal through PLD interaction.