The biarylpyrazole compound AM251 alters mitochondrial physiology via proteolytic degradation of ERRα.

The orphan nuclear receptor estrogen-related receptor alpha (ERR alpha) directs the transcription of nuclear genes involved in energy homeostasis control and the regulation of mitochondrial mass and function. A crucial role for controlling ERR alpha-mediated target gene expression has been ascribed to the biarylpyrazole compound 1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-1-piperidinyl-1H-pyrazole-3-carboxamide (AM251) through direct binding to and destabilization of ERR alpha protein. Here, we provide evidence that structurally related AM251 analogs also have negative impacts on ERR alpha protein levels in a cell-type-dependent manner while having no deleterious actions on ERR gamma. We show that these off-target cellular effects of AM251 are mediated by proteasomal degradation of nuclear ERR alpha. Cell treatment with the nuclear export inhibitor leptomycin B did not prevent AM251-induced destabilization of ERR alpha protein, whereas proteasome inhibition with MG132 stabilized and maintained its DNA-binding function, indicative of ERR alpha being a target of nuclear proteasomal complexes. NativePAGE analysis revealed that ERR alpha formed a similar to 220-kDa multiprotein nuclear complex that was devoid of ERR gamma and the coregulator peroxisome proliferator-activated receptor gamma coactivator-1. AM251 induced SUMO-2,3 incorporation in ERR alpha in conjunction with increased protein kinase C activity, whose activation by phorbol ester also promoted ERR alpha protein loss. Down-regulation of ERR alpha by AM251 or small interfering RNA led to increased mitochondria biogenesis while negatively impacting mitochondrial membrane potential. These results reveal a novel molecular mechanism by which AM251 and related compounds alter mitochondrial physiology through destabilization of ERR alpha.