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Simple Summary: Verticillium dahliae (Vd) is a kind of soil-borne filamentous fungus that can cause Verticillium wilt in many crops and serious damage to agricultural production. Vd 991 is a main pathogen of cotton Verticillium wilt. Previously, we isolated C17 mycosubtilin from Bacillus subtilis J15 (BS J15) broth and showed a significant inhibitory effect on cotton Verticillium wilt. However, the specific fungistatic mechanism by which C17 mycosubtilin antagonizes Vd 991 is not clear. Here, we found that C17 mycosubtilin inhibited the growth and spore germination of Vd 991. C17 mycosubtilin treatment caused shrinking, sinking, and even damage to spores, resulting in rough deformation and uneven distribution of hyphal contents, damage to the cell membrane and cell wall, and mitochondrial swelling of fungi. Moreover, C17 mycosubtilin induced necrosis of Vd 991 in a time-dependent manner. Differential transcription analysis showed that C17 mycosubtilin inhibited fungal growth mainly by destroying the cell structure, blocking the cell cycle, disturbing substance, energy metabolism, and so on. Our results reveal the potential mechanism by which C17 mycosubtilin antagonizes Vd 991, providing useful information for development of more effective antimicrobials. Verticillium wilt is a kind of soil-borne plant fungal disease caused by Verticillium dahliae (Vd). Vd 991 is a strong pathogen causing cotton Verticillium wilt. Previously, we isolated a compound from the secondary metabolites of Bacillus subtilis J15 (BS J15), which showed a significant control effect on cotton Verticillium wilt and was identified as C17 mycosubtilin. However, the specific fungistatic mechanism by which C17 mycosubtilin antagonizes Vd 991 is not clear. Here, we first showed that C17 mycosubtilin inhibits the growth of Vd 991 and affects germination of spores at the minimum inhibitory concentration (MIC). Morphological observation showed that C17 mycosubtilin treatment caused shrinking, sinking, and even damage to spores; the hyphae became twisted and rough, the surface was sunken, and the contents were unevenly distributed, resulting in thinning and damage to the cell membrane and cell wall and swelling of mitochondria of fungi. Flow cytometry analysis with ANNEXINV-FITC/PI staining showed that C17 mycosubtilin induces necrosis of Vd 991 cells in a time-dependent manner. Differential transcription analysis showed that C17 mycosubtilin at a semi-inhibitory concentration (IC50) treated Vd 991 for 2 and 6 h and inhibited fungal growth mainly by destroying synthesis of the fungal cell membrane and cell wall, inhibiting its DNA replication and transcriptional translation process, blocking its cell cycle, destroying fungal energy and substance metabolism, and disrupting the redox process of fungi. These results directly showed the mechanism by which C17 mycosubtilin antagonizes Vd 991, providing clues for the mechanism of action of lipopeptides and useful information for development of more effective antimicrobials. [ABSTRACT FROM AUTHOR]

Simple Summary: Drought and salt stresses are the two major abiotic constraints on plant growth and development, as well as crop production. Previous studies have shown that several of 11 OsOSCAs complement hyperosmolality-induced [Ca2+]cytincreases (OICIcyt), salt stress-induced [Ca2+]cyt increases (SICIcyt), and the associated growth phenotype in the Arabidopsis mutant osca1. However, their biological functions in rice are still unclear. In this paper, we found that OsOSCA1.1 mediates OICIcyt and SICIcyt in rice roots, which are associated with stomatal closure and seedling survival, in response to hyperosmolality and salt stress. The transcriptomic analysis revealed the following three types of OsOSCA1.1-regulated genes in shoots: 2416 sorbitol, 2349 NaCl, and 1844 osmotic-stress response genes. The GO enrichment analysis showed that these OsOSCA1.1-regulated genes were enriched in transcription regulation, hormone responses, and phosphorylation terms, consistent with the enrichment of cis-regulatory elements, i.e., ABRE, ARE, MYB, and MYC binding elements, in the 2000-bp promoter regions of these genes. Our results provide important clues for dissecting calcium regulated gene expression networks in response to drought and salt stresses in rice. OSCA (reduced hyperosmolality-induced [Ca2+]i increase) is a family of mechanosensitive calcium-permeable channels that play a role in osmosensing and stomatal immunity in plants. Oryza sativa has 11 OsOSCA genes; some of these were shown to complement hyperosmolality-induced [Ca2+]cyt increases (OICIcyt), salt stress-induced [Ca2+]cyt increases (SICIcyt), and the associated growth phenotype in the Arabidopsis thaliana mutant osca1. However, their biological functions in rice remain unclear. In this paper, we found that OsOSCA1.1 mediates OICIcyt and SICIcyt in rice roots, which are critical for stomatal closure, plant survival, and gene expression in shoots, in response to hyperosmolality and the salt stress treatment of roots. Compared with wild-type (Zhonghua11, ZH11) plants, OICIcyt and SICIcyt were abolished in the roots of 10-day-old ososca1.1 seedlings, in response to treatment with 250 mM of sorbitol and 100 mM of NaCl, respectively. Moreover, hyperosmolality- and salt stress-induced stomatal closure were also disrupted in a 30-day-old ososca1.1 mutant, resulting in lower stomatal resistance and survival rates than that in ZH11. However, overexpression of OsOSCA1.1 in ososca1.1 complemented stomatal movement and survival, in response to hyperosmolality and salt stress. The transcriptomic analysis further revealed the following three types of OsOSCA1.1-regulated genes in the shoots: 2416 sorbitol-responsive, 2349 NaCl-responsive and 1844 common osmotic stress-responsive genes after treated with 250 mM of sorbitol and 125 mM NaCl of in 30-day-old rice roots for 24 h. The Gene Ontology enrichment analysis showed that these OsOSCA1.1-regulated genes were relatively enriched in transcription regulation, hormone response, and phosphorylation terms of the biological processes category, which is consistent with the Cis-regulatory elements ABRE, ARE, MYB and MYC binding motifs that were overrepresented in 2000-bp promoter regions of these OsOSCA1.1-regulated genes. These results indicate that OsOSCA-mediated calcium signaling specifically regulates gene expression, in response to drought and salt stress in rice. [ABSTRACT FROM AUTHOR]