Expression of Oryza sativa OsSte12 transcription factor influencing antioxidant enzymes, sugar-related operons, and sugar metabolism in Escherichia coli

Authors

  • Kuan-Hung LIN Chinese Culture University, Department of Horticulture and Biotechnology, Taipei, 111 (TW)
  • Chih-Chun CHEN Ming Chuan University, Department of Biotechnology, Taoyuan, 333 (TW)
  • Shwu-Fen PAN Ming Chuan University, Department of Biotechnology, Taoyuan, 333 (TW)
  • Yu-Tsung LEE Chang Gung University of Science and Technology, College of Human Ecology, Research Center for Food and Cosmetic Safety, Taoyuan, 333 (TW)
  • Minh Tan PHAM Ton Duc Thang University, Faculty of Applied Sciences, Ho Chi Minh city, 70000 (VN)
  • Chih-Ming CHIANG Ming Chuan University, Department of Biotechnology, Taoyuan, 333 (TW)

DOI:

https://doi.org/10.15835/nbha53414824

Keywords:

C2H2 zinc fingers, heterologous expressions, lactose, sugar metabolism

Abstract

Ste12 is a C2H2 zinc finger protein transcription factor involved with mating pheromones and regulating protein pathways in microorganisms. Previously, we isolated Ste12 cDNA from rice (Oryza sativa) involved with sugar starvation of α-Amylase in seeds. In this study, we investigated how OsSte12regulated antioxidant enzymes, sugar-related operons, and sugar metabolism by over-expressing OsSte12 in Escherichia coli via transformation. When transformed OsSte12 E. coli was grown in glucose and lactose media, it used these substrates and expressed more activity in ascorbate peroxidase, superoxide dismutase, and sucrose synthase compared to a non-transformant (NT) E. coli strain. Moreover, transformants could be grown in lactose for higher ꞵ-galactosidase activity than NTs or in mannitol. In lactose medium, highly-expressed RNA levels of LacI and LacY were found in transformants, while LacZ gene expression in transformants was significantly reduced compared to NTs. In sucrose and fructose media, FruB and FruK transcripts were both significantly higher in transformants than in NTs, whereas FruA transcripts did not show significant differences between transformants and NTs. Compared to NTs, OsSte12transcriptions of all transformants were significantly up-regulated in response to all sugar sources, but transformants were over-expressed more highly when was grown in lactose than in other sugars. Our results are important for elucidating the regulatory mechanisms of sugars and provide new insights into physiological relevance in OsSte12 transformed E. coli.

References

Aidelberg G, Towbin BD, Rothschild D, Dekel E, Bren A, Alon U (2014). Hierarchy of non-glucose sugars in Escherichia coli. BMC Systtem Biology 8:133. https://doi.org/10.1186/s12918-014-0133-z

Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Research 25(17):3389-3402. https://doi.org/10.1093/nar/25.17.3389

Asunción García-Sánchez M, Martín-Rodrigues N, Ramos B, de Vega-Bartol JJ, Perlin MH, Díaz-Mínguez JM (2010). Fost12, the Fusarium oxysporum homolog of the transcription factor Ste12, is upregulated during plant infection and required for virulence. Fungal Genetetic Biology 47:216-225. https://doi.org/10.1016/j.fgb.2009.11.006

Bernaerts M, De Ley J (1963). A biochemical test for crown gall bacteria. Nature 197:406-407. https://doi.org/10.1038/197406b0

Bouhouche N, Syvanen M, Kado CI (2000). The origin of prokaryotic C2H2 zinc finger regulators. Trends in Microbiology 8(2):77-81. https://doi.org/10.1016/s0966-842x(99)01679-0

Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, … Wittwer CT (2009). The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clinical Chemistry 55(4):611-22. https://doi.org/10.1373/clinchem.2008.112797

D’Abrosca G, Paladino A, Baglivo I, Russo L, Sassano M, Grazioso R, … Malgieri G (2020). Structural insight of the full-length Ros protein: A prototype of the prokaryotic zinc-finger family. Scientific Reports 10(1):9283. https://doi.org/10.1038/s41598-020-66204-5

Diricks M, De Bruyn F, Van Daele P, Walmagh M, Desmet T (2015). Identification of sucrose synthase in nonphotosynthetic bacteria and characterization of the recombinant enzymes. Appllied Microbiology and Biotechnology 99(20):8465-8474. https://doi.org/10.1007/s00253-015-6548-7

Elion E, Qi M, Chen W (2005). Signaling specificity in yeast. Science 307:687-688. https://doi.org/10.1126/science.1109500

Engström-Öst J, Glippa O, Feely RA, Kanerva M, Keister JE, Alin SR, Carter BR, McLaskey AK, Vuori KA, Bednarsek N (2019). Eco-physiological responses of copepods and pteropods to ocean warming and acidification. Scientific Reports 9(1):4748. https://doi.org/10.1038/s41598-019-41213-1

Esposito S, Ilaria B, Gaetano M, Luigi R, Laura Z, Luca D, D'A, Marco M, Benedetto DB, Carla I, Roberto F, Paolo V (2006). A novel type of zinc finger DNA binding domain in the Agrobacterium tumefaciens transcriptional regulator Ros. Biochemistry 45(34):10394-10405. https://doi.org/10.1021/bi060697m

Gao R, Stock AM (2009). Biological insights from structures of two-component proteins. Annual Review of Microbiology 63:133-154. https://doi.org/10.1146/annurev.micro.091208.073214

Garg SG, Hochberg GK (2025). A general substitution matrix for structural phylogenetics, capturing remote homology. Molecular Biology and Evolution 42(6):124. https://doi.org/10.1093/molbev/msaf124

Gomes MP, Kitamura RS, Marques RZ, Barbato ML, Zamocky M (2022). The role of H2O2-scavenging enzymes (ascorbate peroxidase and catalase) in the tolerance of Lemna minor to antibiotics: Implications for phytoremediation. Antioxidants (Basel) 11(1):151. https://doi.org/10.3390/antiox11010151

Gu Q, Zhang C, Liu X, Ma Z (2015). A transcription factor FgSte12 is required for pathogenicity in Fusarium graminearum. Molecular Plant Pathology 16:1-13. https://doi.org/10. 1111/mpp.12155

Gruber S, Zeilinger S (2014). The transcription factor Ste12 mediates the regulatory role of the Tmk1 MAP kinase in mycoparasitismand vegetative hyphal fusion in the filamentous fungus Trichoderma atroviride. PLoS One 9:e111636. https://doi.org/10.1371/journal.pone.0111636

Hamamsy T, Morton JT, Blackwell R, Berenberg D, Carriero N, Gligorijevic V, … Bonneau R (2024). Protein remote homology detection and structural alignment using deep learning. Nature Biotechnology 42(6):975-985. https://doi.org/10.1038/s41587-023-01917-2

Hemmadi V (2016). Metallothionein- A potential biomarker to assess the metal contamination in marine fishes- A review. International Journal of Bioassays 5(4):4961-4973. https://doi.org/10.21746/ijbio.2016.04.003

Hoi WS, Dumas B (2010). Ste12 and Ste12-like proteins, fungal transcription factors regulating development and pathogenicity. Eukaryote Cell 9: 480-485. https://doi.org/10.1128/ec.00333-09

Howlett BJ, Idnurm A, Pedras MS (2001). Leptosphaeria maculans, the causal agent of blackleg disease of Brassicas. Fungal Genetics and Biology 33(1):1-14. https://doi.org/10.1006/fgbi.2001.1274

Janczarek M (2022). The Ros/MucR zinc-finger protein family in bacteria: Structure and functions. International Journal of Molecular Science 23:15536. https://doi.org/10.3390/ijms232415536

Jia H, Zhou Q, Li P, Li M, Li X, Liu Z, Gong X, Dong J, Gu S, Liu Y (2025). Characterization of the C2H2 zinc finger protein family in Setosphaeria turcica. Agronomy 15(6):1434. https://doi.org/10.3390/agronomy15061434

Juers DH, Matthews BW, Huber RE (2012). LacZ β-galactosidase: structure and function of an enzyme of historical and molecular biological importance. Protein Science 21(12):1792-807. https://doi.org/10.1002/pro.2165

Kim TS, Hye YK, Jin HY, Hyen SK (2004a). Recruitment of the Swi/Snf complex by Ste12-Tec1 promotes Flo8-Mss11-mediated activation of STA1 expression. Molecular Cellular Biology 24:9542-9556. https://doi.org/10.1128/MCB.24.21.9542-9556.2004

Kim TS, Sung BL, Hyen SK (2004b). Glucose repression of STA1 expression is mediated by the Nrg1 and Sfl1 repressors and the Srb8-11 complex. Molecular Cellular Biology 24:7695-7706. https://doi.org/10.1128/MCB.24.17.7695-7706.2004

Lang, TJ, Brodsky S, Manadre W, Vidavski M, Valinsky G, Mindel V, Ilan G, Carmi M, Jonas F, Barkai N (2024). Massively parallel binding assay (MPBA) reveals limited transcription factor binding cooperativity, challenging models of specificity. Nucleic Acids Research 52(20):12227-12243. https://doi.org/10.1093/nar/gkae846

Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, … Higgins DG (2007). Cluster W and Cluster X version 2.0. Bioinformatics 23:2947-2948. https://doi.org/10.1093/bioinformatics/btm404

Lewis M (2005) The lac repressor. Comptes Rendus Biologies 328(6):521-548. https://doi.org/10.1016/j.crvi.2005.04.004

Liu JJ, Zhang GC, Oh EJ, Pathanibul P, Turner TL, Jin YS (2016). Lactose fermentation by engineered Saccharomyces cerevisiae capable of fermenting cellobiose. Journal of Biotechnology 234:99-104. https://doi.org/10.1016/j.jbiotec.2016.07.018

Li W, Huang W, Zhou J, Wang J, Liu J, Li Y (2023). Evaluation and control of Alternaria alternata causing leaf spot in soybean in Northeast China. Journal of Applied Microbiology 16:134. https://doi.org/10.1093/jambio/lxad004

Lin KH, Sei SC, Su YH, Chiang CM (2019). Overexpression of the Arabidopsis and winter squash superoxide dismutase genes enhances chilling tolerance via ABA-sensitive transcriptional regulation in transgenic Arabidopsis. Plant Signal Behavior 14(12):1685728. https://doi.org/10.1080/15592324.2019.1685728

Liu L, Ji Z, Zhao K, Zhao Y, Zhang Y, Huang S (2022). Validation of housekeeping genes as internal controls for gene expression studies on biofilm formation in Bacillus velezensis. Applied Microbiology and Biotechnology 106(5):2079-2089. https://doi.org/10.1007/s00253-022-11831-3

Lu CA, Lim EK, Yu SM (1998). Sugar response sequence in the promoter of a rice alpha-amylase gene serves as a transcriptional enhancer. Journal of Biological Chemistry 273:10120-10131. https://doi.org/10.1074/jbc.273.17.10120

Lu CA, Ho T-H D, Ho SL, Yu SM (2002). Three novel MYB proteins with one DNA binding repeat mediate sugar and hormone regulation of α-amylase gene expression. The Plant Cell 14: 1963-1980. https://doi.org/10.1105/tpc.001735

Lu L, Xiao M, Xu X, Li Z, Li Y (2007). A novel beta-galactosidase capable of glycosyl transfer from Enterobacter agglomerans B1. Biochemical and Biophysical Research Communications 27:356(1):78-84. https://doi.org/10.1016/j.bbrc.2007.02.106

Lyu X, Wang Q, Liu A, Liu F, Meng L, Wang P, Zhang Y, Wang L, Li Z, Wang W (2023). The transcription factor Ste12-like increases the mycelial abiotic stress tolerance and regulates the fruiting body development of Flammulina filiformis. Fronteir Microbiology 14:1139679. https://doi.org/10.3389/fmicb.2023.1139679

Madhani HD, Fink GR (1997). Combinatorial control required for the specificity of yeast MAPK signaling. Science 275:1314-1317. https://doi.org/10.1126/science.275.5304.1314

Malgieri G, Luigi R, Sabrina E, Ilaria B, Laura Z, Emilia MP, Benedetto DB, Carla I, Paolo VP, Roberto F (2007). The prokaryotic Cys2His2 zinc-finger adopts a novel fold as revealed by the NMR structure of Agrobacterium tumefaciens Ros DNA-binding domain. PNAS 104:17341-17346. https://doi.org/10.1073/pnas.0706659104

Malgieri G, Palmieri M, Russo L, Fattorusso R, Pedone PV, Isernia C (2015). The prokaryotic zinc‐finger: structure, function and comparison with the eukaryotic counterpart. The FEBS Journal 282(23):4480-4496. https://doi.org/10.1111/febs.13503

Merlini L, Dudin O, Martin SG (2013). Mate and fuse: how yeast cells do it. Open Biology 3:130008. https://doi.org/ 10.1098/rsob.130008

Pinheiro S, Nadal-Ribelles M, Solé C, Vincenzetti V, Dusserre Y, Posas F, Pelet S (2025). Basal association of a transcription factor favors early gene expression. PLoS Genetics 21(6):e1011710. https://doi.org/10.1371/journal.pgen.10117

Record MT, Courtenay DS, Guttman HJ (1998). Responses of E. coli to osmotic stress: large changes in amounts of cytoplasmic solutes and water. TIBS 23 (4):143-148. https://doi.org/10.1016/S0968-0004(98)01196-7

Remmert M, Biegert A, Hauser A, Söding J (2012). HHblits: Lightning-fast iterative protein sequence searching by HMM-HMM alignment. Nature Methods 9(2):173-175. https://doi.org/10.1038/nmeth.1818

Ren R, Yue X, Li J, Xie S, Guo S, Zhang Z (2020). Coexpression of sucrose synthase and the SWEET transporter, which are associated with sugar hydrolysis and transport, respectively, increases the hexose content in Vitis vinifera L. grape berries. Frontier Plant Science 11:321. https://doi.org/10.3389/fpls.2020.00321

Rispail N, Di Pietro A (2010). The homeodomain transcription factor Ste12: Connecting fungal MAPK signaling to plant pathogenicity. Communicative and Integrative Biology 3:327-332. https://doi.org/10.4161/cib.3.4.11908

Schalamun M, Hinterdobler W, Schinnerl J, Brecker L, Schmoll M (2024). The transcription factor STE12 influences growth on several carbon sources and production of dehydroacetic acid (DHAA) in Trichoderma reesei. Scientific Report 14(1):9625. https://doi.org/10.1038/s41598-024-59511-8.5

Schneider CA, Rasband WS, Eliceiri KW (2012). NIH Image to ImageJ: 25 years of image analysis. Natural Methods 9(7):671-5. https://doi.org/10.1038/nmeth.2089

Sezonov G, Joseleau-Petit D, D'Ari R (2007). Escherichia coli physiology in Luria-Bertani broth. Journal of bacteriology 189(23):8746-8749. https://doi.org/10.1128/jb.01368-07

Stock AM, Robinson VL, Goudreau PN (2000). Two-component signal transduction. Annual Review of Biochemistry 69:183-215. https://doi.org/10.1146/annurev.biochem.69.1.183

Subramanian R, Pandi NU, Thangavel R, Swamy LR, Lakshminarayanan SP, Santham S, Ravalan BM, Jacob R (2024). Functional validation of mungbean LEA protein coding gene in bacterial expression system confers salt stress tolerance. Notulae Botanicae Horti Agrobotanici Cluj-Napoca 52(3):13416. https://doi.org/10.15835/nbha52313416

Taylor S, Wakem M, Dijkman G, Alsarraj M, Nguyen M (2010). A practical approach to RT-qPCR-Publishing data that conform to the MIQE guidelines. Methods 50(4):S1-S5. https://doi.org/10.1016/j.ymeth.2010.01.005

Tamura K, Stecher G, Kumar S (2021). MEGA11: Molecular evolutionary genetics analysis Version 11. Molecular Biology 38(7):3022-3027. https://doi.org/10.1093/molbev/msab120

Tan LR, Liu JJ, Deewan A, Lee JW, Xia PF, Rao CV, Jin YS, Wang SG (2022). Genome-wide transcriptional regulation in Saccharomyces cerevisiae in response to carbon dioxide. FEMS Yeast Research 22(1):foac032. https://doi.org/10.1093/femsyr/foac032

Turner TL, Kim E, Hwang C, Zhang GC, Liu JJ, Jin YS (2017). Short communication: Conversion of lactose and whey into lactic acid by engineered yeast. Journal of Dairy Science 100(1):124-128. https://doi.org/10.3168/jds.2016-11784

Vargas WA, Pontis HG, Salerno GL (2008). New insights on sucrose metabolism: evidence for an active A/N-Inv in chloroplasts uncovers a novel component of the intracellular carbon trafficking. Planta 227(4):795-807. https://doi.org/10.1007/s00425-007-0657-1

Wong Sak Hoi J, Dumas B (2010). Ste12 and Ste12-like proteins, fungal transcription factors regulating development and pathogenicity. Eukaryote Cell 9(4):480-485. https://doi.org/10.1128/EC.00333-09

Wei QL, Du YR, Jin K, Xia YX (2017). The Ste12-like transcription factorMaSte12 is involved in pathogenicity by regulating the appressorium formation in the entomopathogenic fungus, Metarhizium acridum. Applied Microbiology and Biotechnology 101:8571–8584. https://doi.org/10.1007/s00253-0178

Woudenberg JH, Seidl MF, Groenewald JZ, de Vries M, Stielow JB, Thomma BP, Crous PW (2015). Alternaria section Alternaria: Species, formae speciales or pathotypes? Study in Mycology 82:1-21. https://doi.org/10.1016/j.simyco.2015.07.001

Yu L, Yang Y, Qiu X, Xiong D, Tian C (2024). The mitogen-activated protein kinase module CcSte11-CcSte7-CcPmk1 regulates pathogenicity via the transcription factor CcSte12 in Cytospora chrysosperma. Stress Biology 4(1):4. https://doi.org/10.1007/s44154-023-00142-w

Yu L, Yang Y, Xiong D, Tian C (2022). Phosphoproteomic and meta bolomic profiling uncovers the roles of CcPmk1 in the pathogenicity of Cytospora chrysosperma. Microbiology Spectrum 10:e0017622. https://doi.org/10. 1128/spect rum. 00176-22

Zhang X, Blumenthal RM, Cheng X (2024). Updated understanding of the protein–DNA recognition code used by C2H2 zinc finger proteins. Current Opinion in Structural Biology 87:102836. https://doi.org/10.1016/j.sbi.2024.102836

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2025-12-21

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LIN, K.-H., CHEN, C.-C., PAN, S.-F., LEE, Y.-T., PHAM, M. T., & CHIANG, C.-M. (2025). Expression of Oryza sativa OsSte12 transcription factor influencing antioxidant enzymes, sugar-related operons, and sugar metabolism in Escherichia coli. Notulae Botanicae Horti Agrobotanici Cluj-Napoca, 53(4), 14824. https://doi.org/10.15835/nbha53414824

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Vol. 53 No. 4 (2025)

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DOI: 10.15835/nbha53414824

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