Vitamin-D (VitD) is proposed to have actions upon skeletal muscle, with supplementation in athletes being shown to improve muscle function, enhance muscle fibre cross sectional area (1), and mitochondrial respiration in human skeletal muscle cells (2). Further to this, higher serum VitD levels have been shown to immediately increase post-exercise, and are associated with improved muscle recovery (3). Additionally, epidemiological studies have linked low serum VitD to impaired skeletal muscle mass/function, and metabolic dysfunction. VitD is known to auto-regulate and act through the vitamin-D receptor (VDR), with VDR expression confirmed in muscle (4). Moreover, in response to resistance exercise, expression of the VDR has been shown to increase in skeletal muscle (5) and this been linked to muscle regeneration and remodeling (4). However, there is currently no defined mechanistic link between VitD and muscle mass/metabolism. In this study, we hypothesized that the VDR has a mechanistic role within the regulation of skeletal muscle mass and metabolism. To probe the mechanistic and muscle-cell autonomous role of the VDR, Tibialis anterior (TA) muscle of Wistar rats were electroporated (under 2.5% isofluorane) to constitutively over-express (VDR-OE) or under-express (VDR-KD) VDR by cDNA and shRNA lenti-viral mediated transfection; contralateral legs were sham treated to act as internal controls. All rats were given carprofen (50 mg/kg) after electroporation and humanely killed before muscles harvested for analysis. VDR-OE yielded myofibre hypertrophy (cross-sectional area (CSA) +17±7%, P<0.05) and increased protein content (+57±12%, P<0.01) compared to contralateral controls. VDR-OE increased both myofibrillar (+44%±12%,P<0.05) and sarcoplasmic (+60%±20%, P<0.01) protein synthesis, with corresponding increases in multiple anabolic signalling protein activities and abundances e.g. mTOR (+93%±30%, P<0.05), RPS6 (+71%±20%, P<0.05) and 4E-BP1 (+57%±21%, P<0.05). Increases in gene expression of several ribosomal proteins namely, RPS11 (+43%±21%, P<0.05), RPS13 (+54%±36%, P<0.05) and RPS28 (+79%±24%, P<0.05) were shown within VDR-OE muscles compared to controls, matching observed increases in total RNA content (+38%±11%, P<0.01, normalised to protein content). Parallel experiments of VDR-KD revealed contrasting effects to VDR-OE: reductions in myofibre CSA (~8±2%, P<0.001) and protein content (~28±16%, P<0.05) compared to sham controls in tandem to markers of increasing autophagy, e.g. protein expression of Cathepsin L (+72±29%, P<0.05) and LC3B-II (+84±43%, P<0.05). The VDR plays a positive role in skeletal muscle mass and metabolism, increasing anabolic capacity, resulting in hypertrophy; in contrast, loss of VDR induces atrophy by up-regulation of autophagy. Our findings are the first to define mechanistic links between the VDR and modulation of in vivo muscle mass and metabolism, perhaps explaining effects of VitD deficiency and supplementation.