The interaction between the measles virus nucleoprotein and the Interferon Regulator Factor 3 relies on a specific cellular environment

BACKGROUND: The genome of measles virus consists of a non-segmented single-stranded RNA molecule of negative polarity, which is encapsidated by the viral nucleoprotein (N) within a helical nucleocapsid. The N protein possesses an intrinsically disordered C-terminal domain (aa 401–525, N(TAIL)) that is exposed at the surface of the viral nucleopcapsid. Thanks to its flexible nature, N(TAIL )interacts with several viral and cellular partners. Among these latter, the Interferon Regulator Factor 3 (IRF-3) has been reported to interact with N, with the interaction having been mapped to the regulatory domain of IRF-3 and to N(TAIL). This interaction was described to lead to the phosphorylation-dependent activation of IRF-3, and to the ensuing activation of the pro-immune cytokine RANTES gene. RESULTS: After confirming the reciprocal ability of IRF-3 and N to be co-immunoprecipitated in 293T cells, we thoroughly investigated the N(TAIL)-IRF-3 interaction using a recombinant, monomeric form of the regulatory domain of IRF-3. Using a large panel of spectroscopic approaches, including circular dichroism, fluorescence spectroscopy, nuclear magnetic resonance and electron paramagnetic resonance spectroscopy, we failed to detect any direct interaction between IRF-3 and either full-length N or N(TAIL )under conditions where these latter interact with the C-terminal X domain of the viral phosphoprotein. Furthermore, such interaction was neither detected in E. coli nor in a yeast two hybrid assay. CONCLUSION: Altogether, these data support the requirement for a specific cellular environment, such as that provided by 293T human cells, for the N(TAIL)-IRF-3 interaction to occur. This dependence from a specific cellular context likely reflects the requirement for a human or mammalian cellular co-factor.