Discovery and characterization of microproteins

Abstract: Recent advances in next-generation sequencing and proteogenomics have revealed thousands of microprotein-encoding small open reading frames in both prokaryotic and eukaryotic genomes, but we are only just beginning to understand their functions. We describe new quantitative and enrichment-based proteomic methods to reveal regulated, stress-responsive translation of unannotated microproteins, as well as molecular validation of non-AUG start codons from novel genes deriving from both prokaryotes and eukaryotes. Finally, we describe functional characterization of the human microprotein NoBody, which inhibits mRNA decay via association with RNA-protein granules called P-bodies and, more specifically, with the protein EDC4. Taken together, our results suggest that quantitative proteomics and molecular characterization can combine to reveal the functions of novel microproteins.