Characterization of Calcium Binding and Coiled Coil Domain 1 (Calcoco1) in Skeletal Muscle
Authors: Irvin Parker , Patterson Kelsey , Waddell David 
Published: June 2021

Skeletal muscle atrophy results in loss of muscle mass and reduced strength and is caused by a range of physiological conditions including aging, cancer, corticosteroid use, and denervation. To further our understanding of the molecular genetic events associated with muscle atrophy, a previous study isolated skeletal muscle from mice following 3 days and 14 days of denervation and the gene expression profile was analyzed by microarray and compared to control muscle in order to identify novel, atrophy‐induced genes. The microarray revealed that Calcium Binding and Coiled Coil Domain 1 (Calcoco1) is expressed in skeletal muscle and induced in response to denervation. The cDNA of Calcoco1 was successfully amplified and cloned from cultured C2C12 cells, demonstrating that this gene is expressed in muscle cells. Quantitative PCR was subsequently conducted using RNA isolated from proliferating and differentiating muscle cells to determine the expression profile of Calcoco1 at the transcriptional level. We observed moderate activation of Calcoco1 through myoblast proliferation and early differentiation, followed by a robust increase in activation by the later stages of myotube differentiation. These results were mirrored at the protein level where it was observed that Calcoco1 is expressed in early proliferation and increases as differentiation progresses. To elucidate Calcoco1's functional role in muscle, we transfected cultured muscle cells with a Calcoco1 expression plasmid and then harvested the cells at timepoints ranging from proliferation through late differentiation. The cell lysates were probed by Western blot for markers of muscle cell differentiation, including Myosin Heavy Chain (MyHC) and myogenin, which both show significant repression in response to Calcoco1 over expression. The Calcoco1 protein is predicted to contain both a calcium binding domain and a coiled‐coiled domain, suggesting that it may play a role in protein‐protein interactions or as a putative transcription factor. To begin to investigate these possibilities, we sought to determine the sub‐cellular localization of Calcoco1 in muscle cells by fusing Calcoco1 cDNA to Green Fluorescent Protein (GFP) and expressing it in muscle cells. Visualization by confocal microscopy revealed clear nuclear‐exclusion of Calcoco1 protein in unchallenged myoblast cells, suggesting that it may not participate directly in gene regulation. The discovery that Calcoco1 is expressed in skeletal muscle and is induced in response to neurogenic atrophy, in combination with its apparent role in muscle cell differentiation, helps further our understanding of the molecular genetic and cellular events of muscle atrophy and may eventually contribute to the identification of new therapeutic targets for the treatment and prevention of muscle wasting.

Support or Funding Information

The work was support by University of North Florida Transformational Learning Opportunity grants and a University of North Florida Foundation Board Grant to D.W.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.