Annexin V Associates with the IFN-γ Receptor and Regulates IFN-γ Signaling

Abstract

Many of the biological activities of IFN-γ are mediated through the IFN-γR3-linked Jak-Stat1α pathway. However, regulation of IFN-γ signaling is not fully understood, and not all responses to IFN-γ are Stat1α dependent. To identify novel elements involved in IFN-γ cell regulation, the cytoplasmic domain of the R2 subunit of the human IFN-γR was used as bait in a yeast two-hybrid screen of a human monocyte cDNA library. This identified annexin A5 (AxV) as a putative IFN-γR binding protein. The interaction was confirmed in pull-down experiments in which a GST-R2 cytoplasmic domain fusion protein was incubated with macrophage lysates. Furthermore, immunoprecipitation using anti-IFN-γR2 Abs showed that AxV interacted with IFN-γR2 to form a stable complex following incubation of cells with IFN-γ. In 293T cells with reduced expression of AxV, brought about by small interfering RNA targeting, activation of Jak2 and Stat1α in response to IFN-γ was enhanced. Inhibition of cell proliferation, a hallmark of the IFN-γ response, also was potentiated in HeLa cells treated with small interfering RNA directed at AxV. Taken together, these results suggest that through an inducible association with the R2 subunit of the IFN-γR, AxV modulates cellular responses to IFN-γ by modulating signaling through the Jak-Stat1 pathway.