Single clone expression cloning and protein microarray platforms for the identification of novel viral immunomodulatory proteins (VIR9P.1157)

Abstract

Abstract Viruses encode proteins that interact with surface receptors on immune cells to evade recognition and elimination by the host immune system. In this study, we focused on the E3/49K protein of human adenoviruses D (HAdV D) a highly diverse protein uniquely found in HAdVs D that was recently demonstrated to suppress leukocyte activation and effector functions through an interaction with receptor tyrosine phosphatase CD45 on immune cells. To advance the search for novel immunomodulatory receptors targeted by the E3/49K protein, we utilized two complementary high-throughput screening technologies consisting of single clone expression cloning and protein microarray platforms. Two highly divergent E3/49 proteins, encoded by HAdV-28 and HAdv-53 serotypes, were systematically probed against thousands of interactions either expressed on the cell-surface or displayed on a protein microarray. We found that both HAdV-28 and HAdV-53 E3/49K proteins interacted with CD45. Furthermore, we report the discovery of two additional receptors specifically targeted by HAdV-28, the leukocyte immunoglobulin-like receptor 1 (LILRB1), a well known immune receptor with inhibitory functions, and putative neuronal cell adhesion molecule (PUNC). Interestingly, we found that PUNC is expressed in immune cells, suggesting a previously undescribed function in immunity. Here we demonstrate the feasibility of this approach for identifying proteins with previously unknown involvement in the human immune response.