F Actin Bundling Dynamics and Stiffness of the TRIOBP-4 F Actin Bundle

Abstract

The guanine nucleotide exchange factor (GEF) trio binding protein isoforms 4 and 5 (TRIOBP-4/-5) is an F-actin-bundling protein originally found to be associated with hearing loss both in humans and mice. The human and mouse TRIOBP protein is largely classified into three isoforms with a long isoform (TRIOBP-5) and two shorter isoforms equivalent to the N-terminus and C-terminus of TRIOBP-5, which are named TRIOBP-4 and TRIOBP-1 (or Tara), respectively. There are no common sequences between TRIOBP-1 and -4. TRIOBP-1 is ubiquitously expressed and has been shown to regulate adherens junctions and actin cytoskeleton reorganization mostly in stress fibers and cortical F-actin. In contrast, TRIOBP-4 and -5 are predominantly expressed in the inner ear and the retina of normal adult tissues, where they are believed to provide appropriate durability and rigidity for stereocilia of hair cells for normal hearing. Interestingly, different from fascin, which is intercalated between actin filaments via its two major actin binding sites, TRIOBP-4 and -5 form extremely dense F-actin bundles without detectable inter-filament spaces, raising the possibility that they more likely wrap around actin bundles. To date, the biological functions of TRIOBP-4/-5 have not been revealed in any other tissues except in inner ear hair cells for hearing. Recently, we identified actin binding sites of TRIOBP-4 isoforms and discovered that TRIOBP-4 expressed in cancer cells to facilitate cell migration speed. In this study, we will show F-actin bundling dynamics during actin polymerization under TRIF microscopy and determine the stiffness and durability of TRIOBP-4/F-actin bundles.