The Cotton Apoplastic Protein CRR1 Stabilizes Chitinase 28 to Facilitate Defense against the Fungal Pathogen <i>Verticillium dahliae</i>

Abstract

The apoplast serves as the first battlefield between the plant hosts and invading microbes; therefore, work on plant-pathogen interactions has increasingly focused on apoplastic immunity. In this study, we identified three proteins in the apoplast of cotton (<i>Gossypium</i> sp) root cells during interaction of the plant with the fungal pathogen <i>Verticillium dahliae</i> Among these proteins, cotton host cells secrete chitinase 28 (Chi28) and the Cys-rich repeat protein 1 (CRR1), while the pathogen releases the protease VdSSEP1. Biochemical analysis demonstrated that VdSSEP1 hydrolyzed Chi28, but CRR1 protected Chi28 from cleavage by <i>Verticillium dahliae</i> secretory Ser protease 1 (VdSSEP1). In accordance with the in vitro results, CRR1 interacted with Chi28 in yeast and plant cells and attenuated the observed decrease in Chi28 level that occurred in the apoplast of plant cells upon pathogen attack. Knockdown of <i>CRR1</i> or <i>Chi28</i> in cotton plants resulted in higher susceptibility to <i>V. dahliae</i> infection, and overexpression of <i>CRR1</i> increased plant resistance to <i>V</i> <i>dahliae</i>, the fungus <i>Botrytis cinerea</i>, and the oomycete <i>Phytophthora parasitica</i> var <i>nicotianae</i> By contrast, knockout of <i>VdSSEP1</i> in <i>V. dahliae</i> destroyed the pathogenicity of this fungus. Together, our results provide compelling evidence for a multilayered interplay of factors in cotton apoplastic immunity.