The non-canonical Smoothened-AMPK axis regulates Smaug1 biomolecular condensates

Biomolecular condensates (BMCs) emerge as important players in RNA regulation. The RNA binding protein Smaug forms cytosolic BMCs in mammals, insects and yeasts and affects mitochondrial function and/or responses to nutrient deprivation. Here we found that the non-canonical activation of the SMO-AMPK pathway known to affect energy metabolism triggers the immediate disassembly of BMCs formed by a number of human and rodent Smaug orthologs, whereas processing bodies remained rather unaltered. A non-phosphorylatable SMO mutant abrogated the effect, involving SMO phosphorylation in hSmaug1 BMCs regulation. Three mechanistically different SMO ligands, namely SAG; GSA-10 and cyclopamine elicited a similar response, which was blocked upon AMPK pharmacological inhibition. Polysome disassembly by puromycin halted Smaug1 BMC dissolution, thus suggesting that unbound transcripts became translationally active. Single-molecule fluorescent in situ hybridization illustrated the release of UQCRC1 mRNA. Finally, Smaug1 is a phosphoprotein bound by 14-3-3 proteins and the competitive inhibitor difopein blocked the response to non-canonical SMO stimulation. We propose that the regulated condensation and dispersion of Smaug1 BMCs generate translational changes that contribute to metabolic regulation downstream of the non-canonical SMO-AMPK axis.