Role of the intracellular domain of the human type I interferon receptor 2chain (IFNAR2c) in interferon signaling - Expression of IFNAR2c truncationmutants in U5A cells

A human cell line (USA) lacking the type I interferon (IFN) receptor chain 2 (IFNAR2c) was used to determine the role of the IFNAR2c cytoplasmic domai n in regulating IFN-dependent STAT activation, interferon-stimulated gene f actor 3 (ISGF3) and c-sis-inducible factor (SIF) complex formation, gene ex pression, and antiproliferative effects, A panel of U5A cells expressing tr uncation mutants of IFNAR2c on their cell surface were generated for study. Janus kinase (JAK) activation was detected in all mutant cell lines; howev er, STAT1 and STATE activation was observed only in U5A cells expressing fu ll-length IFNAR2c and IFNAR2c truncated at residue 462 (R2.462). IFNAR2c mu tants truncated at residues 417 (R2.417) and 346 (R2.346) or IFNAR2c mutant lacking tyrosine residues in its cytoplasmic domain (R2.Y-F) render the re ceptor inactive. A similar pattern was observed for IFN-inducible STAT acti vation, STAT complex formation, and STAT-DNA binding. Consistent with these data, IFN-inducihle gene expression was ablated in U5A, R2.Y-F, R2.417, an d R2.346 cell lines, The implications are that tyrosine phosphorylation and the 462-417 region of IFNAR2c are independently obligatory for receptor ac tivation. In addition, the distal 53 amino acids of the intracellular domai n of IFNAR2c are not required for IFN-receptor mediated STAT activation, IS FG3 or SIF complex formation, induction of gene expression, and inhibition of thymidine incorporation. These data demonstrate for the first time that both tyrosine phosphorylation and a specific domain of IFNAR2c are required in human cells for IFN-dependent coupling of JAK activation to STAT phosph orylation, gene induction, and antiproliferative effects. In addition, huma n and murine cells appear to require different regions of the cytoplasmic d omain of IFNAR2c for regulation of IFN responses.