Myoglobin maturation is driven by the hsp90 chaperone machinery and by soluble guanylyl cyclase

Abstract

Myoglobin (Mb) maturation involves heme incorporation as a final step. We investigated a role for heat shock protein (hsp) 90 in Mb maturation in C2C12 skeletal muscle myoblasts and cell lines. We found the following: 1) Hsp90 directly interacts preferentially with heme-free Mb both in purified form and in cells. 2) Hsp90 drives heme insertion into apoprotein-Mb in an ATP-dependent process. 3) During differentiation of C2C12 myoblasts into myotubes, the apo-Mb-hsp90 complex associates with 5 cell cochaperons, Hsp70, activator of hsp90 ATPase protein 1 (Aha1), alanyl-tRNA synthetase domain containing 1 (Aarsd1), cell division cycle 37 (Cdc37), and stress induced phosphoprotein 1 (STIP1) in a pattern that is consistent with their enabling Mb maturation. 4) Mb heme insertion was significantly increased in cells that had a functional soluble guanylyl cyclase (sGC)-cGMP signaling pathway and was diminished upon small interfering RNA knockdown of sGCβ1 or upon overexpression of a phosphodiesterase to prevent cGMP buildup. Together, our findings suggest that hsp90 works in concert with cochaperons (Hsp70, Aha1, Aarsd1, STIP1, and Cdc37) and an active sGC-cGMP signaling pathway to promote heme insertion into immature apo-Mb, and thus generate functional Mb during muscle myotube formation. This fills gaps in our understanding and suggests new ways to potentially control these processes.—Ghosh, A., Dai, Y., Biswas, P., Stuehr, D. J. Myoglobin maturation is driven by the hsp90 chaperone machinery and by soluble guanylyl cyclase. FASEB J. 33, 9885–9896 (2019). www.fasebj.org.