Brown adipose tissue (BAT) and skeletal muscle expend energy through thermogenesis and manipulation of thermogenesis is a promising target to counteract obesity. Caloric restriction is associated with reduced thermogenesis via homeostatic mechanisms to preserve body mass. We aimed to quantify the combined effects of diet and exercise on thermogenesis in sheep. Ewes (n=5) were treated as follows; 1) ad lib diet, sedentary (AS), 2) ad lib diet, exercise (AE), 3) diet (30% food restriction), sedentary (DS) or 4) diet, exercise (DE). Exercise involved running at 8km/ h for 30 min/day, 5 days/ week, for 4 weeks. Tissue temperature was continuously recorded in sternal fat, retroperitoneal fat and skeletal muscle. There was no effect of exercise on food intake in ad lib-fed animals. Body fat was reduced (P<0.05) in DE sheep only. Night-time (00:00-06:00h) thermogenesis was lowered (P<0.05) in the sternal and retroperitoneal fat of DS sheep. Exercise alone did not affect thermogenesis, but counteracted the effects of food-restriction as observed in DE sheep. There was no effect of diet or exercise on muscle thermogenesis. Protein and gene expression of various markers of thermogenesis were assessed. PSAT-1 (white adipocyte marker) mRNA levels were lower (P<0.05) in the retroperitoneal fat of the DE group, consistent with reduced adiposity. Exercise increased expression of PGC1α (marker of brown and beige adipocytes) in retroperitoneal fat, an effect that was abolished by food restriction. There was no effect of diet or exercise on the expression of uncoupling proteins in fat or muscle. In conclusion, we show that in sheep combined moderate exercise and food-restriction reduces adiposity. Food-restriction causes a compensatory reduction in BAT thermogenesis, which is counteracted by exercise. This is a novel means by which exercise may elicit beneficial metabolic effects.