FGGY Carbohydrate Kinase Domain Containing (Fggy) is Expressed and Alternatively Spliced in Skeletal Muscle

Abstract

Skeletal muscle atrophy can result from a range of physiological conditions, including denervation, corticosteroid exposure, immobilization and aging. In order to better characterize the molecular genetic events of atrophy, a previous investigation isolated skeletal muscle from mice following 3 days and 14 days of denervation. The gene expression profile of the denervated muscle tissue was analyzed by microarray and compared to control muscle in order to identify novel atrophy‐induced genes. The microarray revealed that FGGY carbohydrate kinase domain containing (Fggy) is expressed in skeletal muscle and is induced in response to denervation. Bioinformatic analysis of the Fggy gene locus revealed two validated alternative isoforms, which we have termed Fggy‐L and Fggy‐S, that have distinct transcription initiation sites. In order to confirm that Fggy is expressed in muscle cells, the cDNAs of the two validated alternative splice variants were cloned from mouse myoblast cells. Interestingly, a novel alternative spice variant for each of the validated alternative isoforms was also cloned from mouse muscle cells, suggesting that at least four Fggy variants are expressed in skeletal muscle. In order to better characterize the expression profile of the Fggy gene, quantitative RT‐PCR (RT‐qPCR) was performed using RNA isolated from muscle cells and primer pairs that distinguish all four alternative Fggy transcripts. The qPCR data revealed that the Fggy‐L transcripts are more highly expressed in proliferating muscle cells, while the Fggy‐S transcripts are more evenly expressed in muscle cells that are both proliferating and differentiated. Western Blot analysis of protein homogenates isolated from muscle cells confirmed that the Fggy‐L isoform is more highly expressed in proliferating muscle cells, while the Fggy‐S isoform is more ubiquitously expressed. In addition, confocal fluorescent microscopy analysis showed that the Fggy‐L full length and novel proteins are localized throughout the cell, while the Fggy‐S full length and novel gene products produce a punctuate pattern throughout the cytoplasm of muscle cells. Finally, ectopic expression of Fggy in cultured muscle cells resulted in significant inhibition of muscle cell differentiation. The characterization of novel genes that are activated during neurogenic atrophy helps improve our understanding of the molecular and cellular events that lead to muscle atrophy and could eventually lead to new therapeutic targets for the treatment of muscle wasting.Support or Funding InformationThe 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.