Differential Functions of Two Metalloproteases, Mrmep1 and Mrmep2, in Growth, Sporulation, Cell Wall Integrity, and Virulence in the Filamentous Fungus Metarhizium robertsii

Abstract

The Metarhizium genus of filamentous entomopathogenic fungi plays a pivotal role in regulating insect populations. Metalloproteases (MEPs) are a widely distributed and diverse family of hydrolytic enzymes that are important toxicity factors in the interactions between fungi and their hosts. Herein, we characterized two MEPs, Mrmep1 and Mrmep2, in Metarhizium robertsii using gene deletion. Growth rates of the resulting ΔMrmep1 and ΔMrmep2 mutants decreased by 16.2 and 16.5%, respectively, relative to the wild-type (WT) strain. Both mutants were less sensitive to cell wall-perturbing agents, sodium dodecyl sulfate and Congo red than the WT strain, whereas did not show any obvious changes in fungal sensitivity to ultraviolet B irradiation or heat stress. The conidial yield of ΔMrmep1, ΔMrmep2, and ΔMrmep1ΔMrmep2 mutants decreased by 56.0, 23, and 53%, respectively. Insect bioassay revealed that median lethal time values against Galleria mellonella increased by 25.5% (ΔMrmep1), 19% (ΔMrmep2), and 28.8% (ΔMrmep1ΔMrmep2) compared with the WT strain at a concentration of 1 × 107 conidia mL-1, suggesting attenuated fungal virulence in the ΔMrmep1, ΔMrmep2, and ΔMrmep1ΔMrmep2 strains. During fungal infection, transcription levels of Mrmep1 was 1.6-fold higher than Mrmep2 at 36 h post inoculation. Additionally, transcription levels of gallerimycin gene were 1.2-fold, 2.18-fold, and 2.5-fold higher in insects infected with the ΔMrmep1, ΔMrmep2, or ΔMrmep1ΔMrmep2 mutant than those infected with the WT strain, respectively. Our findings suggest that Mrmep1 and Mrmep2 are differentially contributed to the growth, sporulation, cell wall integrity, and virulence of M. robertsii.

Genome-Wide Identification of M35 Family Metalloproteases in Rhizoctonia cerealis and Functional Analysis of RcMEP2 as a Virulence Factor during the Fungal Infection to Wheat

“…The transcriptional profiles of RcMEP2, RcMEP3, RcMEP4, and RcMEP5 were examined during the infection process to wheat basal sheaths/stems, including five different inoculation time points (18,36,72,96, and 240 h post inoculation (hpi)) and in vitro culturing hypha. Quantitative RT-PCR (qRT-PCR) analyses indicated that during in vitro culturing, RcMEP2, RcMEP3, RcMEP4, and RcMEP5 displayed a very low transcriptional level set as 1, respectively.…”

“…After purification, these His-TF-RcMEP2, His-TF-RcMEP2-1, and His-TF-RcMEP2-2 proteins were checked using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) ( Figure 3C). stems/sheaths, including five different infection time points (18,36,72,96, and 240 h post inoculation (hpi)) and compared to in vitro culturing. Error bars were calculated based on three replicates.…”

“…Metalloproteases include a diverse family of metal ion-dependent proteases and distribute widely in a broad diversity of pathogens [17,18]. Among them, protein sequences of zinc ion-dependent metalloproteases all contain the common HEXXH motif, in which two histidine (H) residues function as the first and the second zinc ligand, respectively [19][20][21].…”

G-protein subunit Gai in mitochondria, MrGPA1, affects conidiation, stress resistance, and virulence of entomopathogenic fungus Metarhizium robertsii

“…To assess the effects of MrGpa1 on virulence, bioassays with Galleria mellonella larvae (RuiQing Bait, Shanghai, China) were performed as described previously (Zhou et al 2018). The larvae were immersed in conidial suspension (1 ´ 10 6 conidia /ml) for 90 s or injected (into the hemocoel) with 10 ml of conidial suspensions (1 ´ 10 5 conidia /ml) and incubated at 25 °C, Each treatment was performed in triplicate, with 18 larvae in each group.…”

Characterization of the active fragments of Spodoptera litura Lebocin‐1

“…Abuse of pesticides have caused serious threats to human health, environment, and the extinction of beneficial species (Gao et al, ; J. Xiao et al, ; J. J. Xiao et al, ). In recent years, more studies have focused on the development of biological approaches to effectively control S. litura (Li et al, ; Liu, Chen, Li, Yang, & Long, ; Wang et al, ; Zhou, Zhou, Fan, Wang, & Huang, ). The immune system of insects plays a crucial role in the success of biological control measures (Luo et al, ; Qian et al, ; Wang et al, ).…”