Adult skeletal muscle fibers show an ability to undergo phenotypic alterations without cell death or regeneration in response to environmental changes. Important factors affecting the metabolic and contractile properties of a muscle fiber includes the activation of genes involved in mitochondrial biogenesis and oxidative phosphorylation, as well as fast and slow isoforms of contractile proteins. The coactivator peroxisome proliferator-activated receptor (PPAR) gamma coactivator (PGC)-1β has recently been proposed to initiate these processes by altering oxygen capacity and myosin heavy chain (MyHC) expression in individual muscle fibers in transgenic animals. However, it is difficult to know if the observed effects reflect a true adult plasticity, or an effect of PGC-1β overexpression throughout myognesis. Here we compared wild type expression patterns of PGC-1β in both fast and slow muscles and investigated the effect of PGC-1β on fiber phenotype in adult mice, where developmental factors are not involved. Expression patterns of the endogenous PGC-1β protein were analyzed by subcellular protein fractionation and Western blotting, while overexpression was studied by electroporating a plasmid encoding Flag-PGC-1β into both the slow oxidative soleus (SOL) and the fast glycolytic extensor digitorum longus (EDL). MyHC fiber type distribution was further analyzed among the transfected fibers, and compared to control fibers within the same muscles. The endogenous PGC-1β protein was found to be expressed 36-fold higher in nuclei from EDL than nuclei from SOL. Overexpression studies in SOL resulted in no MyHC alterations in the PGC-1β-transfected fibers. In EDL an increase in 2x fibers at the expense of 2b fibers was seen when comparing PGC-1β-transfected fibers with the sham-transfected fibers. However, sham transfection in EDL also influenced fiber type, a finding we attribute to selective transfection of fibers with low input resistance. Therefore these findings should be interpreted with caution, and the experiments should be repeated under conditions where sham transfection has no effect.