Identification and Characterization of FGGY Carbohydrate Kinase Domain Containing (Fggy) 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, skeletal muscle was isolated 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‐001 and Fggy‐003, 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 the four alternative Fggy transcripts. The RT‐qPCR data reveals that the Fggy‐001 transcripts are more highly expressed in proliferating muscle cells, while the Fggy‐003 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‐001 isoform is more highly expressed in proliferating muscle cells, while the Fggy‐003 isoform is more ubiquitously expressed. Finally, confocal fluorescent microscopy analysis revealed that the Fggy‐001 full length transcript is localized throughout the cell, while the Fggy‐001 novel alternative transcript only appears to be localized to the cytoplasm. Moreover, the Fggy‐003 novel alternative transcript produces a punctuate localization pattern throughout the cytoplasm of proliferating muscle cells. 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 to D.W.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.