Absorbative and metabolic capacity of the rumen epithelium adapt to different supplies of substrate; however, the nature of this adaptation is poorly characterized. This study sought to determine the effect of different levels of rumen degradable starch and fiber on gene expression in the rumen epithelium. Six yearling, ruminally cannulated Holstein heifers (BW 329 ± 11 kg) were used in a partially replicated Latin Square experiment with 4 treatment diets consisting of high and low rumen degradable starch and NDF arranged factorially. Animals underwent a 3-d diet adaptation period and 14-d diet treatment period. In situ digestibility was assessed by incubating nylon bags of each treatment diet in the rumen from d 10 to 14. Rumen papillae biopsy samples were collected from the ventral sac on d 18 and stored in RNALater at −80°C until RNA isolation. Real-time quantitative reverse transcription PCR was used to measure relative gene abundance. Mixed effect models with fixed effects of starch and fiber and random effects of heifer and period were used for all statistical analyses. In situ results demonstrated the diets had different rates of ruminal starch digestion (4.4%/h vs 7.1%/h; P = 0.05), and different ruminal degradability of NDF (29.9% of dietary NDF vs 38.3%; P = 0.003). There was a significant fiber effect on HSP70 (P = 0.02) and MCT4 (P = 0.04), and a tendency for a fiber effect on MCT1 (P = 0.09). When high ruminally degradable fiber diets were fed, MCT1 and MCT4 expression were reduced, which suggests a negative effect on short-chain fatty acid (SCFA) absorption. Also, AKT1 had a significant starch by fiber interaction (P = 0.049). AKT1 regulates different aspects of metabolism suggesting ruminal nutrient availabilities likely affect basic metabolic function within the rumen epithelium. Reduced expression of SCFA transporters and AKT1 suggests different nutrient degradabilities could impair normal absorbative and metabolic function within the rumen epithelium.