Aim The effects of heat shock transcription factor 1 (HSF1) deficiency on the fibre type composition and the expression level of nuclear factor of activated T cells (NFAT) family members (NFATc1, NFATc2, NFATc3 and NFATc4), phosphorylated glycogen synthase kinase 3α (p‐GSK3α) and p‐GSK3β, microRNA‐208b (miR‐208b), miR‐499 and slow myosin heavy chain (MyHC) mRNAs (Myh7 and Myh7b) of antigravitational soleus muscle in response to unloading with or without reloading were investigated. Methods HSF1‐null and wild‐type mice were subjected to continuous 2‐week hindlimb suspension followed by 2‐ or 4‐week ambulation recovery. Results In wild‐type mice, the relative population of slow type I fibres, the expression level of NFATc2, p‐GSK3 (α and β), miR‐208b, miR‐499 and slow MyHC mRNAs (Myh7 and Myh7b) were all decreased with hindlimb suspension, but recovered after it. Significant interactions between train and time (the relative population of slow type I fibres; P = 0.01, the expression level of NFATc2; P = 0.001, p‐GSKβ; P = 0.009, miR‐208b; P = 0.002, miR‐499; P = 0.04) suggested that these responses were suppressed in HSF1‐null mice. Conclusion HSF1 may be a molecule in the regulation of the expression of slow MyHC as well as miR‐208b, miR‐499, NFATc2 and p‐GSK3 (α and β) in mouse soleus muscle.