An AP-GPCR bigram in Phytophthora capsici is essential for virulence and potentially participates in virulence-related signaling pathways
Aspartic proteases (APs) and G-protein-coupled receptors (GPCRs) are widespread in eukaryotes and play important roles in various cellular processes, including protein modification and cell signaling. In oomycetes, these two functional protein domains are combined into a single protein, forming a so-called bigram. However, the function of these AP-GPCR bigrams remains largely unknown. This study focuses on an AP-GPCR bigram in the devastating oomycete plant pathogen Phytophthora capsici, which is encoded by a single copy gene. Bioinformatic analyses confirmed that PcAPG encodes a membrane-associated protein with an N-terminal signal peptide, a central AP domain, and a C-terminal seven-transmembrane GPCR domain. Expression profiling revealed that PcAPG is constitutively expressed, with a significant upregulation during early host infection (1.5—12 hpi). Knocking out PcAPG using a modified CRISPR/Cas9 system resulted in mutants with only minor mycelial growth defects but severely attenuated virulence, indicating a crucial role for PcAPG in plant-pathogen interaction. Comparative transcriptomic analysis between the wild-type strain and a representative mutant suggests that PcAPG participates in regulating the expression of a suite of putative infection-related genes, potentially via a GPCR-mediated signaling pathway. Notably, in the wild-type strain, 158 genes involved in catabolic processes, such as polysaccharide degradation and hydrolase activities (e.g., alpha-galactosidase activity and arabinan catabolic process), were upregulated during infection, whereas in the mutant, these genes were not upregulated. Based on our findings it is concluded that the AP-GPCR bigram PcAPG is a key virulence factor in P. capsici with a putative role in downstream transcriptional activation of multiple catabolic enzymes thereby facilitating host colonization and invasion. This work unveils an AP-GPCR bigram as a unique signaling module in oomycete pathogens and identifies PcAPG as a promising potential target for novel management strategies against Phytophthora induced diseases.