Significant role of μ-calpain (CANP1) in proliferation/survival of bovine skeletal muscle satellite cells.
Calpains are a family of Ca(2+)-dependent intracellular cysteine proteases, including the ubiquitously expressed μ-calpain (CANP1) and m-calpain (CANP2). The CANP1 has been found to play a central role in postmortem proteolysis and meat tenderization. However, the physiological roles of CANP1 in cattle skeletal satellite cells remain unclear. In this study, three small interference RNA sequences (siRNAs) targeting CANP1 gene were designed and ligated into pSilencer plasmid vector to construct shRNA expression constructs. Suppression of CANP1 in satellite cells was evaluated using these shRNA expressing constructs. Our results revealed that all three siRNAs could downregulate the expression of CANP1. Suppression of CANP1 significantly reduced cell viability in cell proliferation when compared with control cells. We found a crosstalk between CANP1 and caspase systems, particularly suppression of CANP1 resulted in an increase in the expressions of apoptotic caspases such as caspase-3, caspase-6, caspase-7, caspase-8, and caspase-9, as well as heat-shock protein (HSP) systems. Additionally, suppression of CANP1 led to the upregulation of other apoptosis and DNA damage-regulating genes whilst at the same time downregulating proliferation, migration, and differentiation-regulating genes. The results of our findings report for the first time that suppression of CANP1 resulted in the activation of caspase and HSP systems which might in turn regulate apoptosis through the caspase-dependent cell death pathway. This clearly demonstrates the key roles of CANP1 in regulation of cell proliferation and survival.
Department of Animal Science and Biotechnology, Chonbuk National University, Jeonju, 561-756, Republic of Korea.
Hoa
H
Van Ba
Hwang
H
Inho
D000704:Analysis of Variance; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002154:Calpain / Q000378:metabolism*; D020169:Caspases / Q000378:metabolism*; D002417:Cattle; D049109:Cell Proliferation; D002470:Cell Survival; D017931:DNA Primers / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics; D006360:Heat-Shock Proteins / Q000378:metabolism; D046228:Microarray Analysis; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D032496:Satellite Cells, Skeletal Muscle / Q000378:metabolism / Q000502:physiology*
D017931:DNA Primers; D006360:Heat-Shock Proteins; D034741:RNA, Small Interfering; D002154:Calpain; D020169:Caspases; C449002:mu-calpain