Hui Geng

A Novel Sterol Regulatory Element-Binding Protein Gene ( sreA ) Identified in Penicillium digitatum Is Required for Prochloraz Resistance, Full Virulence and erg11 ( cyp51 ) Regulation

Penicillium digitatum is the most destructive postharvest pathogen of citrus fruits, causing fruit decay and economic loss. Additionally, control of the disease is further complicated by the emergence of drug-resistant strains due to the extensive use of triazole antifungal drugs. In this work, an orthologus gene encoding a putative sterol regulatory element-binding protein (SREBP) was identified in the genome of P. digitatum and named sreA. The putative SreA protein contains a conserved domain of unknown function (DUF2014) at its carboxyl terminus and a helix-loop-helix (HLH) leucine zipper DNA binding domain at its amino terminus, domains that are functionally associated with SREBP transcription factors. The deletion of sreA (ΔsreA) in a prochloraz-resistant strain (PdHS-F6) by Agrobacterium tumefaciens-mediated transformation led to increased susceptibility to prochloraz and a significantly lower EC50 value compared with the HS-F6 wild-type or complementation strain (COsreA). A virulence assay showed that the ΔsreA strain was defective in virulence towards citrus fruits, while the complementation of sreA could restore the virulence to a large extent. Further analysis by quantitative real-time PCR demonstrated that prochloraz-induced expression of cyp51A and cyp51B in PdHS-F6 was completely abolished in the ΔsreA strain. These results demonstrate that sreA is a critical transcription factor gene required for prochloraz resistance and full virulence in P. digitatum and is involved in the regulation of cyp51 expression.

Homology Modeling and Screening of New 14α-Demethylase Inhibitor (DMI) Fungicides Based on Optimized Expression of CYP51 from Ustilago maydis in Escherichia coli

Ustilago maydis infection is a serious disease affecting corn crops worldwide. Sterol 14α-demethylase (CYP51) is one of the key enzymes of sterol biosynthesis and an effective target of antifungal drugs. To further study the interaction between CYP51 and drugs and exploit more specific 14α-demethylase inhibitor (DMI) fungicides for U. maydis, in this study homology modeling of CYP51 from U. maydis (UmCYP51) templated as the eukaryotic orthologues (the human CYP51) and screening of new DMI fungicides based on optimized expression were carried out for the first time. In addition, XF-113 and ZST-4 were screened by analyzing the spectral characteristics between the purified UmCYP51-35 and fungicides. These results provide a theoretical basis and new ideas for efficient design and development of new antifungal drugs.

Molecular characterization of a new gammapartitivirus isolated from the citrus-pathogenic fungus Penicillium digitatum

To date, partitiviruses, including gammapartitiviruses, have been extensively studied in various fungal hosts but have not been reported in Penicillium digitatum (also called green mold, the pathogenic fungus infecting citrus). In the present work, we isolated and molecularly characterized a double-stranded RNA (dsRNA) partitivirus from citrus green mold, which we have named “Penicillium digitatum gammapartitivirus 1” (PdGV1). The bisegmented genome of PdGV1 contains two dsRNA segments (dsRNA1 and dsRNA2) with a length of 1795 bp and 1622 bp, respectively. Each of the two genomic dsRNAs contains a single open reading frame encoding a putative RNA-dependent RNA polymerase (RdRp) and a coat protein (CP), respectively. Phylogenetic analysis based on RdRp and CP sequences showed that PdGV1 clustered with mycoviruses belonging to the genus Gammapartitivirus, family Partitiviridae, e.g., Penicillium stoloniferum virus S. The 5’- and 3’-untranslated regions (UTRs) of the PdGV1 genomic dsRNAs both contained unique conserved RNA motifs that have never been found in any other partitivirus. This is the first report of a new gammapartitivirus that infects the citrus-pathogenic fungus P. digitatum.

Fungicides Inhibition Analysis by Molecular Docking and Sensitivity Testing of Penicillium italicum

Blue mold, caused by Penicillium. italicum, is one of the most damaging postharvest diseases of citrus fruit. P. italicum Sterol 14α-demethylase (PiCYP51), an important enzyme in membrance sterol biosynthesis, is a key target of antifungal compounds for citrus disease caused by P. italicum. The three-dimensional structure of PiCYP51 from P. italicum Chinese isolate (HS-1) was constructed through homology modeling basing on the crystal structure of human CYP51. After molecular dynamics (MD) simulation, the refined model was assessed by PROCHECK on the quality. Following evaluation on the reliability was performed by investigating the binding interaction of two commercial sterol 14α-demethylase inhibitors (DMIs) with the enzyme. The binding mode predicted by the molecular docking revealed that the DMIs interacted with PiCYP51 mainly through hydrogen-bonding and hydrophobic interactions. Furthermore, the results were compatible with the detected EC50 values, which were determined as 0.25 and 0.31mg/L for tebuconazole and diniconazole. The binding mode of antifungal agents with PiCYP51 can provide references for DMIs optimization, virtual screening, or de novo antifungal compounds design.

Erratum to: “Overexpression, Homology Modeling and Coenzyme Docking Studies of the Cytochrome P450nor2 from Cylindrocarpon tonkinense” [Molecular Biology 50, 320 (2016)]

N. Lia, b, 2, Y. Z. Zhanga, 2, D. D. Lia, Y. H. Niua, J. Liua, S. X. Lia, Y. Z. Yuana, S. L. Chena, H. Genga, and D. L. Liua, * aHubei Key Laboratory of Genetic Regulation and Integrative Biology, School of Life Science, Central China Normal University, Wuhan, 430079 China bKey Laboratory of Crops with High Quality and Efficient Cultivation and Security Control, Yunnan Higher Education Institutions, College of Life Science and Technology, Honghe University, Mengzi, 661100 China *е-mail: deliliu2013@163.com; ldl@mail.ccnu.edu.cn (Submitted July 15, 2016; accepted for publication July 15, 2016)

Predict the Strong Binding Ability Polypeptide of Human α-Enolase with the HLA-DRB1 * 0401

Human α-enolase (ENO1), an evolutionarily conserved and multifunctional protein, is a target self-antigen of rheumatoid arthritis (RA). Rheumatoid arthritis (RA) is genetically associated with MHC class II molecules, such as DRB1*0101, DRB1*0401 and DRB1*0404 allele. Among these DRB1 alleles, DRB1*0401 show the most correlation with RA. However, strong binding ability polypeptide of ENO1 with HLA-DRB1*0401 is still largely unknown. In this study, we used NetMHCII prediction method to predict the strong binding ability polypeptide with HLA-DRB1*0401. Among the 434 predicted fragment peptide, ENO1129-141: PLYRHIADLAGNS showed strong binding with HLA-DR4 and peptide ENO1281-293 KSFIKDYPVVSIE is the second candidate peptide. Based on these result, we choosed EON1129-141 and EON1281-293 polypeptides to do the molecular modeling, and used the molecular dynamics to optimize the three-dimensional structural model. The molecular dynamics results showed that ENO1129-141: PLYRHIADLAGNS and ENO1281-293: KSFIKDYPVVSIE have strong binding ability with HLA-DR4* 0401. In the shared epitope, both ENO1129-141and ENO1281-293 have the very near distance 3.15Å and 3.10Å with K71 of the β1 chain. The main-chain conformations of ENO1129-141 sit more deeply with β1 chain. All together, results indicated that ENO1129-141 and ENO1281-293 bind strong with HLA-DR4 and would be potential T cell epitopes of human α-enolase that induced RA.