Expression profiling of Bombyx mori gloverin2 gene and its synergistic antifungal effect with cecropin A against Beauveria bassiana.
Gloverin2 is a cationic and glycine-rich antimicrobial peptide whose expression can be induced in fat body of silkworm (Bombyx mori) larvae exposed to bacteria. The purpose of this study is to identify the roles of Bombyx mori gloverin2 (Bmgloverin2) during entomopathogenic fungus Beauveria bassiana infection. Fluorescent quantitative real-time PCR analysis indicated that the relative expression level of Bmgloverin2 gene was up-regulated in the silkworm larvae infected by B. bassiana. The cDNA of Bmgloverin2 was cloned from the silkworm by RT-PCR and the DNA segment of the Bmgloverin2 peptide (without signal peptide sequence) was inserted into pCzn1 expression plasmid and expressed in E. coli ArcticExpress (DE3). SDS-PAGE results revealed that soluble recombinant Bmgloverin2 was successfully expressed and purified. Polyclonal antibody against the Bmgloverin2 was successfully produced with the expressed recombinant protein. Western blot analysis indicated that Bmgloverin2 could be detected in the fat body of silkworm larvae infected with B. bassiana, suggesting that the expression of Bmgloverin2 could be induced by B. bassiana infection in silkworm. Antifungal assays indicated that the Bmgloverin2 had a synergistic antifungal activity with B. mori cecropin A (BmCecA) to entomopathogenic fungus B. bassiana both in vitro and in vivo in the silkworm larvae. This is the first report that Bmgloverin2 exhibits synergistic effect with BmCecA in antifungal activity against B. bassiana. The study demonstrates that Bmgloverin2 is an antifungal protein which plays an important role in synergistic antifungal activity with other antimicrobial peptide in silkworm.
Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, China; Chinese Academy of Agricultural Sciences, Zhenjiang 212018, China; Nursing School, Zhenjiang College, Zhenjiang 212003, China.
Dingding
D
Lü
Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, China; Chinese Academy of Agricultural Sciences, Zhenjiang 212018, China.
Tao
T
Geng
Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, China; Chinese Academy of Agricultural Sciences, Zhenjiang 212018, China.
Chengxiang
C
Hou
Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, China; Chinese Academy of Agricultural Sciences, Zhenjiang 212018, China.
Guangxing
G
Qin
Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, China; Chinese Academy of Agricultural Sciences, Zhenjiang 212018, China.
Kun
K
Gao
Sericultural Research Institute, Jiangsu University of Science and Technology, Zhenjiang 212018, China; Chinese Academy of Agricultural Sciences, Zhenjiang 212018, China. Electronic address: guoxijie@126.com.
Xijie
X
Guo
D000818:Animals; D000935:Antifungal Agents / Q000008:administration & dosage*; D023181:Antimicrobial Cationic Peptides / Q000008:administration & dosage* / Q000502:physiology; D052982:Beauveria / Q000187:drug effects* / Q000472:pathogenicity*; D012831:Bombyx / Q000235:genetics* / Q000382:microbiology / Q000502:physiology; D004357:Drug Synergism; D017344:Genes, Insect; D019476:Insect Proteins / Q000008:administration & dosage* / Q000235:genetics* / Q000502:physiology; D036341:Intercellular Signaling Peptides and Proteins; D011506:Proteins / Q000008:administration & dosage* / Q000235:genetics* / Q000502:physiology; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics; D015854:Up-Regulation
D000935:Antifungal Agents; D023181:Antimicrobial Cationic Peptides; D019476:Insect Proteins; D036341:Intercellular Signaling Peptides and Proteins; D011506:Proteins; D011994:Recombinant Proteins; C107617:gloverin; C031161:cecropin A
Beauveria bassiana; BmCecA; Bmgloverin2; Bombyx mori; Synergistic antifungal activity