Here the hypothesis that skeletal muscle Ca²⁺-calmodulin dependent kinase II (CaMKII) expression and signalling would be modified by endurance training was tested. Eight healthy, young men completed 3 wk of one-legged endurance exercise training with muscle samples taken from both legs before training and 15h after the last exercise bout. Along with an ~40% increase in mitochondrial F₁-ATP synthase expression, there was an ~1-fold increase in maximal CaMKII activity and CaMKII kinase isoform expression after training in the active leg only. Autonomous CaMKII activity and CaMKII autophosphorylation were increased to a similar extent. However, there was no change in -CaMKII anchoring protein expression with training. Nor was there any change in expression or Thr¹⁷ phosphorylation of the CaMKII substrate phospholamban with training. However, another CaMKII substrate serum response factor (SRF), had an ~60% higher phosphorylation at Ser¹⁰³ after training, with no change in SRF expression. There were positive correlations between the increases in CaMKII expression and SRF phosphorylation as well as F₁ATPase expression with training. After training, there was an increase in cyclic-AMP response element binding protein phosphorylation at Ser¹³³, but not expression, in muscle of both legs. Taken together, skeletal muscle CaMKII kinase isoform expression and SRF phosphorylation is higher with endurance-type exercise training, adaptations that are restricted to active muscle. This may contribute to greater Ca²⁺ mediated regulation during exercise and the altered muscle phenotype with training.