Characterization of a Neo-mode of Profilin Mediated Formin Regulation

Das, Saikat (2026) Characterization of a Neo-mode of Profilin Mediated Formin Regulation. PhD thesis, Indian Institute of Science Education and Research Kolkata.

Text (PhD thesis of Saikat Das (19RS033)) 19RS033.pdf - Submitted Version Restricted to Repository staff only Download (9MB)

Abstract

The Wnt signaling pathway, conserved across metazoans, regulates embryonic development, comprises of β-catenin-dependent (canonical) and β-catenin-independent (non-canonical) branches. In the planar cell polarity pathway, Wnt binds Frizzled (Fz) receptors, activating Dishevelled2 (Dvl2), which interacts with the formin-binding protein Daam1. In the Wnt/Ca²⁺ pathway, activated Dvl2 triggers phospholipase C (PLC)-mediated intracellular Ca²⁺ release. Daam1, a formin family protein, contains conserved FH1 and FH2 domains; the FH2 domain promotes linear actin filament formation, while the proline-rich FH1 domain binds profilin—a 15 kDa protein that delivers actin monomers to the filament’s barbed end—enhancing formin-mediated elongation. Thus, profilin acts as a formin accelerator. Previous studies show that profilin knockdown in T24M cells suppresses the Wnt/Ca²⁺ pathway, while its knockdown during early development disrupts the non-canonical pathway. This led us to hypothesize that profilin may act upstream of Daam1 to regulate its activity. Our bioinformatic analysis suggests Dvl2 adopts an autoinhibited conformation via intramolecular PDZ–C-terminal interactions. We show that profilin binds to the C-terminal region, potentially relieving Dvl2 autoinhibition and promoting Daam1 interaction. A recent study showed formin-binding protein 1 (FNBP1) interacts with Daam1 and regulates non-canonical Wnt signaling. We recently published that formin-binding protein 4 (FNBP4) regulates Formin1 (FMN1) by binding its WW1 domain to FH1, inhibiting actin nucleation and acting as a decelerator, contrasting profilin. Here, we show that FNBP4 similarly regulates mDia1 and Daam1 via WW1–FH1 interactions, identifying FNBP4 as a pan-formin regulator. Additionally, we found FNBP4’s WW2 domain interacts with profilin, suggesting profilin may also regulate formins indirectly via FNBP4. Altogether, our findings uncover a neo-mode of profilin-mediated formin regulation, wherein profilin modulates formin activity through adaptor proteins such as FNBP4 and Dvl2.

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