Erythropoietin (EPO) signals via its receptor (EPOR) to exert widespread endocrine (erythropoiesis) and paracrine-autocrine (proliferation, apoptosis, angiogenesis, organogenesis, cyto-protection, repair, and carcinogenesis) actions. Progress in EPOR research has been hampered by distinct gaps in knowledge and tools: A) Uncertainty regarding the existence of "antisense EPOR polypeptides". B) Paucity of molecules (agonists or antagonists) binding to EPOR other than EPO itself. C) Incomplete knowledge of binding partners of EPOR through protein-protein interaction. We propose to propel the EPOR field forward by employing focused, short-term, intense efforts that close these gaps using conventional and novel approaches to generate high impact reagents and fundamental data that will be applicable to multiple disciplines. Specific Aims are to: 1) Follow up on the discovery of natural complementary antisense EPOR (asEPOR) transcripts that synergistically enhance EPOR protein expression induced by the sense EPOR mRNA, by determining whether asEPOR transcripts are translated into "antisense polypeptides" in vivo. 2) Identify peptoids that bind to, activate or inactivate EPOR using existing recombinatorial peptoid libraries;peptoid agonists and antagonists can serve as lead compounds for discovery of therapeutic agents. 3) Identify EPOR- binding membrane and cytoplasmic proteins using a novel approach where full length EPOR serves as bait. Identifying EPOR binding partners will open new horizons for manipulating EPOR signaling pathways. These Aims involve 3 investigators and 2 collaborating laboratories with different expertise and resources. With intense work in 2 years, we aim to deliver new reagents and knowledge, and generate resources for the entire EPOR research community. Reagents include 1) If in vivo asEPOR polypeptide is proven, specific antisera will be available. 2) EPOR-modifying peptoids to spark therapeutic drug discovery and for manipulating EPOR signaling in animal models. Knowledge includes 1) Proof or disproof of in vivo asEPOR polypeptides. Proof of asEPOR polypeptides will jumpstart a novel area of research, while disproof will focus future work on regulatory mechanisms of asEPOR transcripts. 2) A complete list of transmembrane and cytoplasmic EPOR binding proteins will be identified and validated to facilitate the study of EPOR signaling. Our approach is discovery in nature rather than hypothesis testing. PUBLIC HEALTH RELEVANCE: Erythropoietin receptor biology impacts basic biomedical as well as clinical disciplines. There is high potential for clinical therapeutic development;hence unequivocal public health benefit to having these results in the public domain. Knowledge and reagents from this project integrate immediately with other NIH funded research.