Ribonuclease T2 from Aspergillus fumigatus promotes T helper type 2 responses through M2 polarization of macrophages

Abstract

Allergic bronchopulmonary aspergillosis (ABPA) is an allergic immunological response to Aspergillus fumigatus (Af) exposure, which induces a strong T helper 2 (Th2) response via mechanisms that have yet to be elucidated. The aim of the present study was to investigate the hypothesis that T2 ribonuclease from Af (Af RNASET2) induces M2-type macrophage polarization to produce a T helper 2 (Th2) immune response. Recombinant Af RNASET2 (rAf RNASET2) was expressed and purified in a prokaryotic pET system and BALB/c mice were immunized with rAf RNASET2 for in vivo analyses. Expression levels of M2 polarization factors were evaluated in RAW264.7 macrophages treated with rAf RNASET2 in vitro using flow cytometry, reverse transcription-quantitative PCR, and western blot analysis. The results predicted that the mature Af RNASET2 protein (382 amino acids; GenBank no. MN593022) contained two conserved amino acid sequence (cAS) domains, termed cAS-1 and cAS-2, which are also characteristic of the RNASET2 family proteins. The protein expression levels of the Th2-related cytokines interleukin (IL)-4, IL-10, and IL-13 were upregulated in mice immunized with rAf RNASET2. RAW264.7 macrophages treated with rAf RNASET2 showed increased mRNA expression levels of M2 factors [arginase 1, Il-10, and Il-13]; however, there was no difference in cells treated with rAf RNASET2 that had been inactivated with a ribonuclease inhibitor (RNasin). The protein expression levels of IL-10 in macrophage culture supernatant were also increased following stimulation with rAf RNASET2. In addition, rAf RNASET2 upregulated the expression of phosphorylated mitogen activated protein kinases (MAPKs) in RAW264.7 cells, whereas MAPK inhibitors attenuated rAf RNASET2-induced IL-10 expression in RAW264.7 cells. In conclusion, the present study reveals that high rAf RNASET2 activity is required for rAf RNASET2-induced M2 polarization of macrophages and suggests an important immune regulatory role for Af RNASET2 in ABPA pathogenesis.

Two Secretory T2 RNases Act as Cytotoxic Factors Contributing to the Virulence of an Insect Fungal Pathogen

“…28,29 The enzyme in A. fumigatus apparently modulates the human immune response. 27 Our results demonstrated that two secretory RNase T2 members of the animal (insect) fungal pathogen, B. bassiana, contribute to virulence primarily via their cytotoxicity to host cells, as well as activation of insect humoral immune responses that depend on their RNases enzyme activities and glycosylation modification. However, the details of those enzymes entering host cells, as well as reaching the plant nucleus from fungal pathogens, are still unclear and need to be revealed in the future work.…”

“…fumigatus secrete RNase T2 members during infection that are required for full fungal virulence, which are seemly due to their enzyme activities in utilization of plant RNA presenting in plant apoplast, cytoplasm, and nucleus. , The enzyme in A. fumigatus apparently modulates the human immune response . Our results demonstrated that two secretory RNase T2 members of the animal (insect) fungal pathogen, B.…”

“…The eggs secrete an RNase T2 family member (termed Omega-1 or SjCP1412) to drive polarization of the host Th2 immune response that aids in the egg’s excretion and transmission to a molluscan intermediate host. , An emerging bacterial pathogen Acinetobacter baumannii secretes an RNase T2 family member that is involved in motility and the ability of the pathogen to colonize inanimate surfaces . The human fungal pathogen Aspergillus fumigatus produces an RNase T2 during infection, which acts as an immune regulatory factor in allergic bronchopulmonary aspergillosis (ABPA), an allergic immunological response, primarily through augmentation of M2 macrophages polarization and thus induction of a Th2 immune response . In the plant fungal pathogen Ustilago maydis, two T2 ribonucleases Nuc1 and Nuc2 are secreted during infection and contribute to fungal virulence, as well as the completion of the pathogenic lifecycle.…”

“…26 The human fungal pathogen Aspergillus fumigatus produces an RNase T2 during infection, which acts as an immune regulatory factor in allergic bronchopulmonary aspergillosis (ABPA), an allergic immunological response, primarily through augmentation of M2 macrophages polarization and thus induction of a Th2 immune response. 27 In the plant fungal pathogen Ustilago maydis, two T2 ribonucleases Nuc1 and Nuc2 are secreted during infection and contribute to fungal virulence, as well as the completion of the pathogenic lifecycle. These effects may be due to their enzyme function in utilization of the extracellular RNA (presenting in maize apoplast) as a phosphate source at least.…”

Two secretory T2 RNases contribute to virulence of an insect fungal pathogen due to cytotoxicity and activation of host immune response

“…Several fungal pathogens including the phytopathogens U. maydis and F. oxysporum, and the human pathogen A. fumigatus secrete RNase T2 during infection that are required for full fungal virulence, which are seemly due to their enzyme activities in utilization of plant RNA presenting in plant apoplast, cytoplasm and nucleus(Mukherjee et al, 2020;Qian et al, 2022). The enzyme in A. fumigatus apparently modulates the human immune response(Chen et al, 2020). Our results demonstrated that two secretory RNase T2 members of the animal (insect) fungal pathogen, B. bassiana, contribute to virulence primarily via their cytotoxicity to host cells, as well as activation of insect humoral immune responses that depend on their RNases enzyme activities and glycosylation modi cation.…”