Protein kinase C-θ (PKC-θ) is a lipid-sensitive molecule associated with lipid-induced insulin resistance in skeletal muscle. Rodent models have not cohesively supported that PKC-θ impairs insulin responsiveness in skeletal muscle. The purpose of this study was to generate mice that lack PKC-θ in skeletal muscle and determine how lipid accumulation and insulin responsiveness are affected in that tissue. Mice lacking PKC-θ in skeletal muscle (SkM^PKCθKO) and controls (SkM^PKCθWT) were placed on a regular diet (RD) or high-fat diet (HFD) for 15 wk, followed by determination of food intake, fasting glucose levels, lipid accumulation, and insulin responsiveness. There were no differences between SkM^PKCθWT and SkM^PKCθKO mice on a RD. SkM^PKCθKO mice on a HFD gained less weight from 10 through 15 wk of dietary intervention (P < 0.05). This was likely due to less caloric consumption (P = 0.0183) and fewer calories from fat (P < 0.001) compared with SkM^PKCθWT mice on a HFD. Intramyocellular lipid accumulation (P < 0.0001), fatty acid binding protein 4, and TNF-α mRNA levels (P < 0.05) were markedly reduced in SkM^PKCθKO compared with SkM^PKCθWT mice on a HFD. As a result, fasting hyperglycemia was mitigated and insulin responsiveness, as indicated by Akt phosphorylation, was maintained in SkM^PKCθKO on a HFD. Liver lipid accumulation was not affected by genotype, suggesting the deletion of PKC-θ from skeletal muscle has a tissue-specific effect. PKC-θ is a regulator of lipid-induced insulin resistance in skeletal muscle. However, the effects of this mutation may be tissue specific. Further work is warranted to comprehensively evaluated whole body metabolic responses in this model.