Functional characterization of the chaperon-like protein Cdc48 in cryptogein-induced immune response in tobacco

Cdc48, a molecular chaperone conserved in different kingdoms, is a member of the AAA+ family contributing to numerous processes in mammals including proteins quality control and degradation, vesicular trafficking, autophagy and immunity. The functions of Cdc48 plant orthologues are less understood. We previously reported that Cdc48 is regulated by S-nitrosylation in tobacco cells undergoing an immune response triggered by cryptogein, an elicitin produced by the oomycete Phytophthora cryptogea. Here, we inv estigated the function of NtCdc48 in cryptogein signalling and induced hypersensitive-like cell death. NtCdc48 was found to accumulate in elicited cells at both the protein and transcript levels. Interestingly, only a small proportion of the overall NtCdc48 population appeared to be S-nitrosylated. Using gel filtration in native conditions, we confirmed that NtCdc48 was present in its hexameric active form. An immunoprecipitation-based strategy following my mass spectrometry analysis led to the identification of about a hundred NtCdc48 partners and underlined its contribution in cellular processes including targeting of ubiquitylated proteins for proteasome-dependent degradation, subcellular trafficking and redox regulation. Finally, the analysis of cryptogein-induced events in NtCdc48-overexpressing cells highlighted a correlation between NtCdc48 expression and hypersensitive cell death. Altogether, this study identified NtCdc48 as a component of cryptogein signalling and plant immunity. In a previous study, we demonstrated that the chaperone-like protein NtCdc48 is regulated by S-nitrosylation in tobacco cells undergoing an immune response triggered by cryptogein, an elicitin produced by the oomycete Phytophthora cryptogea. In the present original study, we further investigated the function of this chaperone in cryptogein signalling and cryptogein-triggered hypersensitive-like cell death. We reported that only a small proportion of the overall NtCdc48 protein population undergoes S-nitrosylation. However, using different strategies such as gel filtration in native conditions, immunoprecipitation and the generation of a tobacco cell line overexpressing NtCdc48, we demonstrated that this protein is mobilized in response to cryptogein, is present as a hexameric and active complex and interacts with numerous partners related to proteasome-dependent degradation, subcellular trafficking and redox regulation. Importantly, our study highlighted a role for NtCdc48 in cryptogein-triggered cell death. Altogether, this investigation designs NtCdc48 as a new component of plant immunity.