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Original Article

Korean J Physiol Pharmacol 2019; 23(6): 539-547

Published online November 1, 2019 https://doi.org/10.4196/kjpp.2019.23.6.539

Copyright © The Korean Journal of Physiology & Pharmacology.

Deficiency of Anoctamin 5/TMEM16E causes nuclear positioning defect and impairs Ca2+ signaling of differentiated C2C12 myotubes

Tam Thi Thanh Phuong#, Jieun An#, Sun Hwa Park, Ami Kim, Hyun Bin Choi, and Tong Mook Kang*

Department of Physiology, Sungkyunkwan University School of Medicine, Suwon 16419, Korea

Correspondence to:Tong Mook Kang
E-mail: tongmkang@skku.edu

#These authors contributed equally to this work.

Received: September 23, 2019; Revised: September 30, 2019; Accepted: September 30, 2019

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.


Anoctamin 5 (ANO5)/TMEM16E belongs to a member of the ANO/TMEM16 family member of anion channels. However, it is a matter of debate whether ANO5 functions as a genuine plasma membrane chloride channel. It has been recognized that mutations in the ANO5 gene cause many skeletal muscle diseases such as limb girdle muscular dystrophy type 2L (LGMD2L) and Miyoshi muscular dystrophy type 3 (MMD3) in human. However, the molecular mechanisms of the skeletal myopathies caused by ANO5 defects are poorly understood. To understand the role of ANO5 in skeletal muscle development and function, we silenced the ANO5 gene in C2C12 myoblasts and evaluated whether it impairs myogenesis and myotube function. ANO5 knockdown (ANO5-KD) by shRNA resulted in clustered or aggregated nuclei at the body of myotubes without affecting differentiation or myotube formation. Nuclear positioning defect of ANO5-KD myotubes was accompanied with reduced expression of Kif5b protein, a kinesin-related motor protein that controls nuclear transport during myogenesis. ANO5-KD impaired depolarization-induced [Ca2+]i transient and reduced sarcoplasmic reticulum (SR) Ca2+ storage. ANO5-KD resulted in reduced protein expression of the dihydropyridine receptor (DHPR) and SR Ca2+-ATPase subtype 1. In addition, ANO5-KD compromised co-localization between DHPR and ryanodine receptor subtype 1. It is concluded that ANO5-KD causes nuclear positioning defect by reduction of Kif5b expression, and compromises Ca2+ signaling by downregulating the expression of DHPR and SERCA proteins.

Keywords: Anoctamin 5, C2C12, Excitation-contraction coupling, Kif5b, Myoblast differentiation, Nuclear positioning,