Polypeptides encoded by short open reading frames (sPEPs) are translated abundantly in prokaryotic and eukarytic cells, yet their functions remain mysterious (Andrews and Rothnagel, 2014). While examples of such peptides performing essential functions in human tissues are known for over a decade, the breadth and relevance of peptides encoded in short open reading frames is only beginning to be explored. Recent, rigorous genome wide prediction and mass spectrometric detection indicates the presence of well over a thousand distinct sPEPs in mammalian cells. Exemplified by a handful of studies characterizing single sPEPs, many of the detected sPEPs will have relevance to human biology and disease. The lack of immunological reagents for sPEPs is the major obstacle in understanding sPEP function.
I will leverage a new method to express proteins with a minimal, unnatural amino acid based, bio-orthogonal handle to perform an in vitro screen for DNA aptamers recognizing sPEP sequences. DNA aptamers are recognized as promising alternative to antibodies for in situ detection, microscopy, histology and diagnostics, due to the enormous diversity that can be generated in in vitro screens, relatively smaller size and high specificity. Using the same technology, a high-throughput approach for characterizing sPEPs function in human cell lines will be developed, assessing subcellular localization and protein interactomes.