Human cytomegalovirus IE1 promoter/enhancer drives variable gene expression in all fiber types in transgenic mouse skeletal muscle

Abstract

Background: Versatile transgenic manipulation of skeletal muscle requires knowledge of the expression profiles of diverse promoter/enhancer elements in the transcriptionally specialized fiber types of which muscle is composed. "Universal" viral promoters/enhancers, e.g., cytomegalovirus IE1 (CMV IE1), are of interest as reagents that may drive broad expression. However, a previous study noted a marked heterogeneity of CMV IE1-driven transgene expression among muscle fibers, raising the possibility of fiber-type-restricted expression. The purpose of the present study was to characterize CMV IE1-driven expression in terms of fiber type. Results:We produced two lines of transgenic mice carrying the CMV IE1/ β-galactosidase construct CMVLacZ, and analyzed transgene expression and fiber type by histochemical analysis of hindlimb muscle sections. In both lines CMVLacZ was expressed in all four major fiber types: type I (slow) and types IIA, IIB and IIX (fast). There was no unique pattern of fiber-type-preferential expression; fiber-type quantitative differences were observed but details varied between muscle regions and between lines. Both lines showed similar fiber-type-independent regional differences in overall expression levels, and a high level of within-fiber-type variability of expression, even among nearby fibers. The soleus muscle showed strong expression and comparatively little within-fibertype or between-fiber-type variability. Conclusions:The CMV IE1 promoter/enhancer is not fiber-type-restricted and can be useful for driving germ-line transgene expression in all four fiber types. However, not all fibers express the gene at high levels due in part to regional differences in overall expression levels, and to a high level of within-fiber-type variability. Given the multinucleate syncitial nature of muscle fibers, it is not likely that this variability is due to variegating heterochromatinization. The soleus muscle would make a suitable subject for near-uniform experimental gene expression driven by CMV IE1 elements.

Local and systemic administration of AAV vectors with alphaherpesvirus latency-associated promoter 2 drives potent transgene expression in mouse liver, kidney, and skeletal muscle

“…Interestingly, earlier studies featuring AAV9-CMV revealed higher rates of transduction of the TA muscle compared to SOL, suggesting that myofiber types may be differentially permissive to AAV-mediated transduction and transgene expression ( Bostick et al, 2007b ). Results from other previous studies suggest that CMV promoters do not discriminate between myofiber types, ( Hallauer and Hastings, 2000 ; Robinson et al, 2005 ) given that SOL muscle consists mainly of type 1 (slow-twitch) fibers, whereas TA, EDL, GA, and QUAD muscles include more type 2 (fast-twitch) fibers ( Komiya et al, 2017 ). Here, we observed that the fluorescence intensity and biodistribution of mCherry resulting from the IM inoculation of AAV8-LAP2 and AAV9-LAP2 drive robust transgene expression.…”

Affinity Sensing for Transgenes Detection in Antidoping Control

“…EGFP gene detection was addressed for its wide use as marker of transgenosis event, both in in vitro and in vivo preclinic trials, and thus represents a well-known reporter system. On the other hand, studies on mice have demonstrated the importance of the CMV promoter in high level and constitutive gene expression, i.e., in skeletal muscle. , …”

Differential myofiber-type transduction preference of adeno-associated virus serotypes 6 and 9

“…In our cassette, GFP was expressed under CMV promoter; thus, we cannot exclude the possibility of suboptimal activity of CMV promoter in MyHC-2b myofibers. However, previous studies showed that the CMV promoter does not discriminate between myofiber types [ 28 , 29 ]. Additional studies with different promoters and reporters could clarify this potential concern.…”