The co-chaperone Cdc37 regulates the rabies virus phosphoprotein stability by targeting to Hsp90AA1 machinery.

Yunbin Xu,Jie Zan,Senlin Ji,Yan Yan,Jiyong Zhou,Fei Liu,Dandan Wang,Juan Liu

doi:10.1038/srep27123

Abstract

Cdc37, as a kinase-specific co-chaperone of the chaperone Hsp90AA1 (Hsp90), actively aids with the maturation, stabilization and activation of the cellular or viral kinase/kinase-like targets. Phosphoprotein (P) of rabies virus (RABV) is a multifunctional, non-kinase protein involved in interferon antagonism, viral transcription and replication. Here, we demonstrated that the RABV non-kinase P is chaperoned by Cdc37 and Hsp90 during infection. We found that Cdc37 and Hsp90 affect the RABV life cycle directly. Activity inhibition and knockdown of Cdc37 and Hsp90 increased the instability of the viral P protein. Overexpression of Cdc37 and Hsp90 maintained P’s stability but did not increase the yield of infectious RABV virions. We further demonstrated that the non-enzymatic polymerase cofactor P protein of all the genotypes of lyssaviruses is a target of the Cdc37/Hsp90 complex. Cdc37, phosphorylated or unphosphorylated on Ser13, aids the P protein to load onto the Hsp90 machinery, with or without Cdc37 binding to Hsp90. However, the interaction between Cdc37 and Hsp90 appears to have additional allosteric regulation of the conformational switch of Hsp90. Our study highlighted a novel mechanism in which Cdc37/Hsp90 chaperones a non-kinase target, which has significant implications for designing therapeutic targets against Rabies.

Highlights

Probed with rabbit anti-ß-actin and anti-GAPDH polyclonal antibody (pAb) and mouse monoclonal antibody (mAb) to N and P to detect the expression of N and P respectively. (E) N2a cells were transfected with Cdc[37] short hairpin RNA (shRNA) or Hsp[90] shRNA for 24 h, and infected with Rabies virus (RABV) strain HEP-Flury at an multiplicities of infection (MOI) = 1 for 36 h
The accumulation of Cdc[37], the Hsp[90] co-chaperone, was markedly promoted after RABV infection at both 12 hpi and 24 hpi (P < 0.001), before Hsp[90] upregulation. These results indicated that RABV infection upregulated the expression of both Cdc[37] and Hsp[90]
Up to date, studies have reported that the chaperone Hsp[90] is nearly universally required for viral protein stability[30], only one kinase and one kinase-like protein of DNA viruses have been shown to interact with Cdc37/Hsp[90] machinery

Summary

Introduction

A Cdc[37] mutant defective in Hsp[90] binding functioned in a dominant-negative fashion by preventing the interaction between Hsp[90] and kinases[18,19,20]. Inhibitors, such as celastrol, lead to target degradation by disruption of Cdc37/Hsp[90] complexes, without interfering with ATP binding to Hsp9021,22. Unlike the chaperoning of the kinase targets, phosphorylation of Cdc[37] is not required for P protein stabilization. Our study highlighted a novel mechanism whereby Cdc37/Hsp[90] chaperones a non-kinase target

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Conclusion