The carboxyl‐terminal ahnak domain induces actin bundling and stabilizes muscle contraction

Abstract

Ahnak, a 700 kDa protein, is expressed in a variety of cells and has been implicated in different cell-type-specific functions. In the human heart, we observed an endogenous carboxyl-terminal 72 kDa ahnak fragment that copurified with myofibrillar proteins. Immunocytochemistry combined with confocal microscopy localized this fragment to the intercalated discs and close to the Z-line of cardiomyocytes. No endogenous myofibrillar ahnak fragment was observed in the skeletal muscle. We elucidated the role of the recombinant carboxyl-terminal ahnak fragment (ahnak-C2) in actin filament organization and in the function of muscle fibers. Addition of ahnak-C2 to actin filaments induced filament bundling into paracrystalline-like structures as revealed by electron microscopy. Incubation of demembranated skeletal muscle fibers with ahnak-C2 attenuated the decline in isometric force development upon repeated contraction-relaxation cycles. Our results suggest that the carboxyl-terminal ahnak domain exerts a stabilizing effect on muscle contractility via its interaction with actin of thin filaments.

Structure of a C-terminal AHNAK peptide in a 1:2:2 complex with S100A10 and an acetylated N-terminal peptide of annexin A2

“…The C-terminus of AHNAK has been shown to be responsible for translocation between nucleus and cytoplasm (Nie et al, 2000) and the induction of actin bundling (Haase et al, 2004), and AHNAK is a major component of enlargosomes, which are vesicles for rapid calcium-dependent exocytosis (Borgonovo et al, 2002). The C-terminal domain of AHNAK interacts with (p11) 2 (AnxA2) 2 and all three proteins co-localize at the intracellular face of the plasma membrane in a Ca 2+ -dependent manner.…”

The C type natriuretic peptide receptor tethers AHNAK1 at the plasma membrane to potentiate arachidonic acid-induced calcium mobilization

“…The carboxy terminal domain of AHNAK1 contains nuclear export and localization signals (16,44) and is responsible for its translocation and subcellular localization (30). AHNAK1 has been implicated in numerous cell-type-specific functions including T cell calcium signaling (24,25), regulation of calcium channel gating properties in cardiomyocytes (1), modulation of DNA ligase IV-mediated double-stranded ligation (43), stabilization of skeletal muscle contractility (17), skeletal muscle regeneration (18), and plasma membrane differentiation and repair (4).…”

Protective Function of Ahnak1 in Vascular Healing after Wire Injury

“…To examine the expression and localization of Ahnak1 and Ahnak2 in murine femoral arteries, we employed previously characterized antibodies developed against the specific carboxy-terminal ends of Ahnak1 and Ahnak2 [5,32]. Confocal microscopy of immunostained sections of murine femoral artery showed prominent expression of Ahnak1 in vascular endothelium (Fig.…”

“…The antibodies against Ahnak1 and Ahnak2 were raised in rabbits using synthetic peptides corresponding to specific sequences in the carboxy-terminal portion of murine large Ahnak1 (L-Ahnak NP_033773.1; KKSRLSSSSSNDSG) and Ahnak2 (AHNAK2 XP_006516522.1; AHNAK2 isoform X1 XP_006543088.1 and X2 XP_006543089.1; GSEAMVASSARTEL), respectively. The antibodies were affinity-purified on the respective peptides, and have been described elsewhere [5,32]. TUNEL assay (in situ cell death detection kit, Roche) was done according to the manufacturer's protocol.…”

“…It is highly expressed in muscle as well as in epithelial and endothelial cell layers with barrier function [4,5,6,7]. First described as desmoyokin in bovine epithelia [8], it was later identified as a tumor-related protein with nuclear localization in cancer cell lines [9,10].…”