EFFECTS OF CA2+ ON LEIOMODIN-2 FUNCTION

Madison Little

The striated muscle thin filament, comprised of a filamentous actin (F-actin) backbone, tropomyosin (Tm) and troponin (Tn), is a fundamental part of Ca2+-dependent muscle contraction, and its length must be maintained for effective contraction. In cardiac muscle, the length of the thin filament is regulated at the pointed end by related proteins tropomodulin-1 (Tmod1) and leiomodin-2 (Lmod2). Tmod1 tightly caps the pointed end to prevent actin polymerization and depolymerization, whereas Lmod2 promotes slow filament elongation. The mechanism governing Lmod2 function and its competition with Tmod1 at the pointed end remain poorly understood. Lmod2 shares an N-terminal tropomyosin-binding site (TmBS) and two actin-binding sites (ABS1 and ABS2) with Tmod1 but is distinguished by a linker between ABS1 and ABS2 suggested to bind Ca2+, and a C-terminal extension of ~20 kDa that houses a third ABS, a Wiskott Aldrich Homology (WH2) domain. The WH2 domain is important for Lmod2 function, and its removal converts Lmod2 into a pointed-end cap resembling Tmod1. Aside from the WH2 domain, the role of the C-terminal extension is unknown. Capping by Lmod2’s N-terminal domains is less efficient than Tmod1, giving rise to the proposal that Lmod2 acts as a leaky cap, however the lack of structural data leaves the molecular basis of this behavior unresolved. This dissertation investigates role of Lmod2’s C-terminal extension and the structural basis of the leaky cap mechanism in pointed end elongation. We identified two novel actin-binding regions in the C-terminal extension and demonstrate that it mediates Ca2+-dependent binding to the side of the thin filament. We further show Ca2+-dependent localization of Lmod2 to the pointed end, together supporting a role of Ca2+ in regulating Lmod2’s association with the thin filament. Finally, we present the structure of the Lmod2 ABS1-actin interface, that provides molecular details of the leaky cap mechanism and explains how Lmod2 and Tmod1 achieve different pointed end functions despite their conserved capping domains. These findings contribute to the mechanistic understanding of thin filament length regulation and reveal Ca2+ as a regulator of Lmod2’s thin filament binding.

EFFECTS OF CA2+ ON LEIOMODIN-2 FUNCTION

Files and links (1)

Abstract

Metrics

Details