*2.2. TRIOBP-1 as a Regulator of Actin Polymerization*

Upon its initial discovery, TRIOBP-1 was noted to adopt a filamentous expression pattern, appearing at 350 nm periodic intervals along the length of actin filaments [3]. Direct interaction between TRIOBP-1 and actin could be demonstrated in vitro, strongly indicating TRIOBP-1 to be an actin-associated protein [3]. Furthermore, TRIOBP-1 co-localizes with, although seemingly does not bind to, two other actin-associated proteins, actinin and myosin II [3]. Knockdown of TRIOBP-1 by siRNA has repeatedly been shown to lower the expression of filamentous F-actin in cell systems [24–26], while over-expression of TRIOBP-1 in cell lines leads to a "cell spreading" phenotype, resulting from excessive F-actin formation [3]. Notably, the central CC domain of TRIOBP-1 is capable of interacting with F-actin and blocking its depolymerization into G-actin [12]. One of the principle cellular functions of TRIOBP-1 therefore appears to be maintaining the existence of F-actin fibers.

In wound healing assays, performed in neuroblastoma cells, overexpression of TRIOBP-1 was seen to increase the rate of cellular migration [19]. This effect was cumulative with that of overexpressing NDEL1 (Nuclear Distribution Element-Like 1, also known as Nudel) [19]. NDEL1 is a key neurodevelopmental protein with links to mental illness, which is more commonly associated with the microtubule cytoskeleton [27]. Nevertheless, NDEL1 directly interacted with TRIOBP-1, binding to the central CC region at approximately the fourth coiled coil, and appeared to work co-operatively with TRIOBP-1 to enhance levels of F-actin [19]. Furthermore, in neuronal systems, TRIOBP-1 appears to recruit two key kinases to NDEL1 [28]. The ensuing phosphorylation events lead to increased F-actin formation, neurite outgrowth, and dendritic arborization [28]. TRIOBP-1 and NDEL1 therefore appear to act synergistically in cell migration and neuronal differentiation.

Another important role of actin is in relation to the receptors that modulate adhesion between the cell and both its extracellular matrix and other cells. The actin cytoskeleton physically links these and provides the basis of mechanical force within the cell that allows it to interact with external stimuli [29]. TRIOBP-1 has been identified in the focal adhesions that link cells to the extracellular matrix, and its expression there is regulated by myosin II [30], which generates tension, leading to maturation of the focal adhesions. TRIOBP-1 is also found at the adhesion junctions between cells [13]. In adhesion junctions of epithelial cells, expression of the crucial transmembrane protein E-cadherin is regulated by the RhoGEF TRIO. TRIOBP-1 binds to TRIO using its mid domain and prevents this effect, leading to increased E-cadherin expression and increased density of actin filaments [13]. It remains to be clarified whether this role of TRIOBP-1 in modulating actin via TRIO is distinct from its effect on actin depolymerization, which seems to occur through direct binding [3,12].

TRIOBP-1 is also found at the adherens junctions in the heart, where it interacts with JCAD (Junctional Protein Associated with Coronary Artery Disease) [31]. Knockdown of either TRIOBP-1 or JCAD in epithelial cells led to reduced F-actin stress fiber formation [31]. TRIOBP-1 also possesses an additional function in the heart through its interaction with the voltage gated ion channel hERG1 (human Ether-à-go-go-Related Gene 1, also known as KCNH2) [15]. In cardiomyocytes, TRIOBP-1 affects expression of hERG, with direct effects on cardiac rapidity, leading the authors to speculate that TRIOBP-1 may function as a bridge between actin filaments and hERG1 in the membrane, linking excitation of the ion channel to cell mobility [15].
