Mutation of key amino acids in the polygalacturonase‐inhibiting proteins CkPGIP1 and GhPGIP1 improves resistance to Verticillium wilt in cotton

Abstract

Verticillium wilt, one of the most devastating diseases of cotton (Gossypium hirsutum), causes severe yield and quality losses. Given the effectiveness of plant polygalacturonase-inhibiting proteins (PGIPs) in reducing fungal polygalacturonase (PG) activity, it is necessary to uncover the key functional amino acids to enhance cotton resistance to Verticillium dahliae. To identify novel antifungal proteins, the selectivity of key amino acids was investigated by screening against a panel of relevant PG-binding residues. Based on the obtained results, homologous models of the mutants were established. The docking models showed that hydrogen bonds and structural changes in the convex face in the conserved portion of leucine-rich repeats (LRRs) may be essential for enhanced recognition of PG. Additionally, we successfully constructed Cynanchum komarovii PGIP1 (CkPGIP1) mutants Asp176Val, Pro249Gln, and Asp176Val/Pro249Gln and G. hirsutum PGIP1 (GhPGIP1) mutants Glu169Val, Phe242Gln, and Glu169Val/Phe242Gln with site-directed mutagenesis. The proteins of interest can effectively inhibit VdPG1 activity and V. dahliae mycelial growth in a dose-dependent manner. Importantly, mutants that overproduced PGIP in Arabidopsis and cotton showed enhanced resistance to V. dahliae, with reduced Verticillium-associated chlorosis and wilting. Furthermore, the lignin content was measured in mutant-overexpressing plants, and the results showed enhanced lignification of the xylem, which blocked the spread of V. dahliae. Thus, using site-directed mutagenesis assays, we showed that mutations in CkPGIP1 and GhPGIP1 give rise to PGIP versatility, which allows evolving recognition specificities for PG and is required to promote Verticillium resistance in cotton by restricting the growth of invasive fungal pathogens.

Transcriptome sequencing of Verticillium dahliae from a cotton farm reveals positive correlation between virulence and tolerance of sugar-induced hyperosmosis

“…However, these small molecules greatly improved the osmotic pressure of the intercellular liquid. Therefore, improving the effectiveness of plant polygalacturonase-inhibiting proteins (PGIPs) in reducing fungal polygalacturonase (PG) activity could enhance cotton resistance to V. dahliae (Liu et al, 2018; Liu et al, 2017b). The winter embolism recovery of the walnut tree, where air bubbles in xylem vessels are cleared by an osmotic force produced via high-level PIP2 expression and the increase of sucrose from the conversion of starch-to-sugar corroborated this (Sakr et al, 2003).…”

GhPLP2 Positively Regulates Cotton Resistance to Verticillium Wilt by Modulating Fatty Acid Accumulation and Jasmonic Acid Signaling Pathway

“…Two weeks after VIGS, when the true leaves of GhCLA1 -silenced cotton plants showed clear signs of albinism, the plants were inoculated with V. dahliae (1 × 10 7 conidia/mL) as previously described ( Liu et al, 2018 ). The plant disease index (DI) was monitored using the following formula as described by Wang et al (2020) : DI = [Σ (n × the number of seedlings at level n)/(4 × the number of total seedlings)] × 100, n denotes disease level, cotton seedlings were divided into five levels based on their disease severity after inoculation with V. dahliae (level 0, 1, 2, 3, 4).…”

Peer Review #2 of "Transcriptome sequencing of Verticillium dahliae from a cotton farm reveals positive correlation between virulence and tolerance of sugar-induced hyperosmosis (v0.1)"

“…However, these small molecules greatly improved the osmotic pressure of the intercellular liquid. Therefore, improving the effectiveness of plant polygalacturonase-inhibiting proteins (PGIPs) in reducing fungal polygalacturonase (PG) activity could enhance cotton resistance to V. dahliae (Liu et al, 2018;Liu et al, 2017b) Manuscript to be reviewed corroborated this (Sakr et al, 2003). Likewise, the V.dahliae could affect the xylem vessels of host plants to produce cavitations and starch hydrolysis, and vessel occlusion was considered relevant to wilt symptoms (Trapero et al, 2018).…”