Presentation Information
[P02-292]Functional analysis of factors involved in ManR-dependent β-1,4-mannobiose induction in Aspergillus
○Konomi Shiba1, Yui Shiga1, Tomoko Shintani2, Shoki Fujita2, Katsuya Gomi2, Shuji Tani1 (1. Graduate School of Agriculture, Osaka Metropolitan University (Japan), 2. Graduate School of Agriculture, Tohoku University (Japan))
Keywords:
Gene Regulation,Filamentous fungi,Cellulase,Enzyme production
[Purpose] In Aspergillus aculeatus, a cellulase-producing filamentous fungus, the expression of cellulase and β-1,4-mannanase genes is regulated by a balance between induction by cellobiose and β-1,4-mannobiose and repression by glucose. We previously demonstrated that the transcription factor ManR mediates the induction of these enzyme genes in response to both inducers. In this ManR-dependent gene induction pathway, we found that disruption of the UDP-glucose 4-epimerase gene (Δuge5) abolished β-1,4-mannobiose-responsive gene induction, resulting in a marked decrease in β-1,4-mannanase production.1 In contrast, deletion of SepL (ΔsepL), which encodes a protein kinase component of the Septation Initiation Network (SIN), abolished cellobiose-responsive induction.2 The A. aculeatus ΔsepL strain exhibited defects in septation and conidiation, as well as increased sensitivity to Congo red, consistent with phenotypes reported in other Aspergillus species. These results suggest that SepL, in addition to its conserved roles, is involved in cellobiose-responsive gene expression in A. aculeatus. To elucidate the molecular mechanisms underlying ManR-dependent responses to distinct inducers, this study focused on the β-1,4-mannobiose-responsive regulatory pathway mediated by UDP-glucose 4-epimerase (Uge).
[Method] A. aculeatus possesses two genes encoding Uge, namely uge3 and uge5. We constructed a single deletion mutant of uge3 (Δuge3) and a double deletion mutant of uge3 and uge5 (Δuge3Δuge5), and quantitatively analyzed their responses to β-1,4-mannobiose.
[Results] The inducible gene expression in response to β-1,4-mannobiose was completely abolished in Δuge5. In Δuge3, their inducible expression was reduced, and the expression level in the presence of 10 mM glucose together with 4 mM β-1,4-mannobiose (repressing conditions) decreased to basal levels comparable to that of the host strain. Unexpectedly, the Δuge3Δuge5 strain under both inducing and repressing conditions exhibited gene expression levels comparable to the host strain under the inducing condition. In humans, G protein-coupled receptors (GPCRs) are involved in regulating cellular processes mediated by nucleotide sugars such as UDP-glucose and UDP-galactose. To investigate the possible involvement of GPCRs in the Uge-dependent β-1,4-mannobiose response, we assessed the effect of the GPCR deletions on enzyme production using a GPCR deletion mutant library in Aspergillus oryzae. Strains were cultured on media containing locust bean gum or carboxymethyl cellulose as the sole carbon source and glutamine, glutamic acid, or diammonium tartrate as the sole nitrogen source.
[Consideration] In this presentation, we will discuss the Uge-mediated mechanism of β-1,4-mannobiose-responsive induction, incorporating results from our ongoing GPCR analyses.
[Conclusion] The unexpected restoration of gene expression in Δuge3Δuge5 mutant suggests the presence of an alternative regulatory mechanism, potentially involving GPCR-mediated signaling. These findings provide new insights into the molecular basis of inducer-specific regulation within the ManR-dependent pathway.
1. Kuga et al. Appl Microbiol Biotechnol. 2023; 107(2-3):785-795; 2. Sawada at al. J Basic Microbiol. 2025; 65(5):e2400266
[Method] A. aculeatus possesses two genes encoding Uge, namely uge3 and uge5. We constructed a single deletion mutant of uge3 (Δuge3) and a double deletion mutant of uge3 and uge5 (Δuge3Δuge5), and quantitatively analyzed their responses to β-1,4-mannobiose.
[Results] The inducible gene expression in response to β-1,4-mannobiose was completely abolished in Δuge5. In Δuge3, their inducible expression was reduced, and the expression level in the presence of 10 mM glucose together with 4 mM β-1,4-mannobiose (repressing conditions) decreased to basal levels comparable to that of the host strain. Unexpectedly, the Δuge3Δuge5 strain under both inducing and repressing conditions exhibited gene expression levels comparable to the host strain under the inducing condition. In humans, G protein-coupled receptors (GPCRs) are involved in regulating cellular processes mediated by nucleotide sugars such as UDP-glucose and UDP-galactose. To investigate the possible involvement of GPCRs in the Uge-dependent β-1,4-mannobiose response, we assessed the effect of the GPCR deletions on enzyme production using a GPCR deletion mutant library in Aspergillus oryzae. Strains were cultured on media containing locust bean gum or carboxymethyl cellulose as the sole carbon source and glutamine, glutamic acid, or diammonium tartrate as the sole nitrogen source.
[Consideration] In this presentation, we will discuss the Uge-mediated mechanism of β-1,4-mannobiose-responsive induction, incorporating results from our ongoing GPCR analyses.
[Conclusion] The unexpected restoration of gene expression in Δuge3Δuge5 mutant suggests the presence of an alternative regulatory mechanism, potentially involving GPCR-mediated signaling. These findings provide new insights into the molecular basis of inducer-specific regulation within the ManR-dependent pathway.
1. Kuga et al. Appl Microbiol Biotechnol. 2023; 107(2-3):785-795; 2. Sawada at al. J Basic Microbiol. 2025; 65(5):e2400266
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