Hsp90 is the ATP-consuming core component of a very abundant molecular chaperone machine that handles a substantial portion of the cytosolic proteome. Rather than one machine, it is in fact an ensemble of molecular machines since most mammalian cells express two cytosolic isoforms of Hsp90, a subset of up to 40-50 co-chaperones, and regulate their interactions and functions by a variety of posttranslational modifications. We demonstrate that the Hsp90 ensemble is fundamentally remodeled during muscle differentiation and that this remodeling is not just a consequence of muscle differentiation, but possibly one of the drivers to accompany and to match the vast proteomic changes associated with this process. As myoblasts differentiate into myotubes, Hsp90α disappears and only Hsp90β remains, the only isoform capable of interacting with the novel muscle-specific Hsp90 co-chaperone Aarsd1L. Artificially maintaining Hsp90α or knocking down Aarsd1L expression interfere with the differentiation of C2C12 myotubes. During muscle differentiation, Aarsd1L replaces the more ubiquitous co-chaperone p23 and in doing so dampens the activity of the glucocorticoid receptor, one of the Hsp90 clients relevant to muscle functions. This co-chaperone switch protects muscle cells against the inhibitory effects of glucocorticoids and may contribute to preventing muscle wasting induced by excess glucocorticoids.