Identification of an essential regulator controlling the production of raw-starch-digesting glucoamylase in Penicillium oxalicum

Abstract

BackgroundRaw-starch-digesting glucoamylases (RSDGs) from filamentous fungi have great commercial values in starch processing; however, the regulatory mechanisms associated with their production in filamentous fungi remain unknown. Penicillium oxalicum HP7-1 isolated by our laboratory secretes RSDG with suitable properties but at low production levels. Here, we screened and identified novel regulators of RSDG gene expression in P. oxalicum through transcriptional profiling and genetic analyses.ResultsPenicillium oxalicum HP7-1 transcriptomes in the presence of glucose and starch, respectively, used as the sole carbon source were comparatively analyzed, resulting in screening of 23 candidate genes regulating the expression of RSDG genes. Following deletion of 15 of the candidate genes in the parental P. oxalicum strain ∆PoxKu70, enzymatic assays revealed five mutants exhibiting significant reduction in the production of raw-starch-digesting enzymes (RSDEs). The deleted genes (POX01907, POX03446, POX06509, POX07078, and POX09752), were the first report to regulate RSDE production of P. oxalicum. Further analysis revealed that ∆POX01907 lost the most RSDE production (83.4%), and that POX01907 regulated the expression of major amylase genes, including the RSDG gene POX01356/PoxGA15A, a glucoamylase gene POX02412, and the α-amylase gene POX09352/Amy13A, during the late-stage growth of P. oxalicum.ConclusionOur results revealed a novel essential regulatory gene POX01907 encoding a transcription factor in controlling the production of RSDE, regulating the expression of an important RSDG gene POX01356/PoxGA15A, in P. oxalicum. These results provide insight into the regulatory mechanism of fungal amylolytic enzyme production.Electronic supplementary materialThe online version of this article (10.1186/s13068-018-1345-z) contains supplementary material, which is available to authorized users.

Gene Regulatory Networks of Penicillium echinulatum 2HH and Penicillium oxalicum 114-2 Inferred by a Computational Biology Approach

“…In N. crassa, the female fertility-7 (FF-7) is required for initiation of sexual development, and ff-7 mutant does not produce protoperithecia and perithecia as well as ascospores (Carrillo et al, 2017). In P. oxalicum HP7-1, it was demonstrated that POX09752, ortholog of PDE_06843 in P. oxalicum 114-2, is involved in the regulation of raw-starchdigesting enzymes (Zhang et al, 2019). This is consistent with the inferred regulatory interactions where PDE_06843 regulates one α-amylase encoding gene PDE_04683 (GH13-5) in P. oxalicum; and PECH_007045 regulates two α-amylase encoding genes PECH_000987 (GH13-5) and PECH_000986 (GH13-1) in P. echinulatum.…”

Constructing a novel expression system by specific activation of amylase expression pathway in Penicillium

“…Generally, glycoside hydrolases expression is regulated by upstream transcription factor(s) in filamentous fungi [ 16 , 21 , 22 ]. Comparative analysis of significantly changed genes in all three strains revealed 40 DEGs encoding putative transcription factors (TFs) (Additional file 8: Table S8), most of which contained zinc-related structures (Zn_clus, GATA, Zn2Cys6, bZIP, etc.).…”

“…Therefore, the dramatic increase in the transcription levels of amy15A benefited from not only the absence of the repressor CreA but also the increase in the activator AmyR. In view of this conclusion, it is desirable to further improve the efficiency of amy15A(p) by overexpressing AmyR or other potential activitors when inhibition effect mediated by CreA is relieved [22]. In addition, the negative regulation of CreA on hyphal growth and enhanced secretion of Amy15A in liquid indicate that Δ13A-OamyR-ΔCreA has higher expression efficiency per unit biomass, which would benefit protein productivity in high-density fermentation.…”

“…P. oxalicum produces large amounts of glycoside hydrolases, most of their production is tightly controlled by complex regulatory networks [ 16 , 29 ]. Besides AmyR and CreA, other novel regulators of amylolytic enzymes were screened and identified in P. oxalicum through transcriptional profiling and genetic analysis [ 22 ]. The differentially expressed TFs in response to the AmyR overexpression and CreA deletion indicated these TFs might directly or indirectly regulate the efficiency of amy15A (p) in this study (Fig.…”

Identification of a Novel Transcription Factor TP05746 Involved in Regulating the Production of Plant-Biomass-Degrading Enzymes in Talaromyces pinophilus

“…Plant-biomass-degrading enzymes activities, including SSDE, raw starch-digesting enzymes (RSDE), cellulase and xylanase activities, were assayed as previously described Zhang et al, 2019). Briefly, 50 µL of appropriately diluted crude extract produced from the parental strain TpKu70 or its deletion mutant TP05746, were added to 450 µL of 100 mM citrate-phosphate buffer (pH 5.0) containing 1% SCS (Sigma-Aldrich), 1% raw cassava flour (farmer's market, Nanning, China), 1% CMC-Na (Sigma-Aldrich, Darmstadt, Germany), or 1% beechwood xylan (Megazyme International Ireland, Bray, Ireland), and 1 mL of 100 mM citrate-phosphate buffer pH 5.0, containing Whatman No.…”