Skip to main content
SpringerLink
Log in

C-terminal Ser/Thr residues are vital for the regulatory role of Ste7 in the asexual cycle and virulence of Beauveria bassiana

  • Biotechnologically relevant enzymes and proteins
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

The mitogen-activated protein kinase (MAPK) kinase Ste7 has a conserved Ser/Thr loop (S/T-X4(6)-S/T) that can activate the MAPK Fus3 or Kss1 for the regulation of pheromone response and filamentous growth in model yeast. Here, we show that not only the loop but also four C-terminal Ser/Thr residues are essential for Ste7 to function in the Fus3 cascade of Beauveria bassiana, a filamentous fungal insect pathogen. Mutagenesis of either looped S216/T220 or C-terminal S362 resulted in the same severe defects in conidial germination, hyphal growth, aerial conidiation, and submerged blastospore production as the ste7 deletion, followed by a complete loss of virulence and similarly increased cell sensitivities to osmotic salts, oxidants, heat shock and UV-B irradiation. Mutagenesis of three other Ser/Thr residues (S391, S440, and T485) also caused severe defects in most of the mentioned phenotypes. These defects correlated well with dramatically reduced transcript levels of some phenotype-related genes. These genes encode a transcription factor (CreA) essential for carbon/nitrogen assimilation, developmental activators (BrlA, AbaA, and WetA) and upstream transcription factor (FluG) required for conidiation, P-type N+/K+ ATPases (Ena1–5) required for intracellular N+/K+ homeostasis, and antioxidant enzymes involved in multiple stress responses. Our study unveils that the loop and four C-terminal Ser/Thr residues are all vital for the regulatory role of Ste7 in the growth, conidiation, virulence, and/or stress tolerance of B. bassiana and perhaps other filamentous fungi.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  • Andrews DL, Egan JD, Mayorga ME, Gold SE (2000) The Ustilago maydis ubc4 and ubc5 genes encode members of a MAP kinase cascade required for filamentous growth. Mol Plant-Microbe Interact 13:781–786

    Article  PubMed  CAS  Google Scholar 

  • Banuett F, Herskowitz I (1994) Identification of fuz7, a Ustilago maydis MEK/MAPKK homolog required for a-locus-dependent and -independent steps in the fungal life cycle. Genes Dev 8:1367–1378

    Article  PubMed  CAS  Google Scholar 

  • Bardwell AJ, Flatauer LJ, Matsukuma K, Thorner J, Bardwell L (2001) A conserved docking site in MEKs mediates high-affinity binding to MAP kinases and cooperates with a scaffold protein to enhance signal transmission. J Biol Chem 276:10374–10386

    Article  PubMed  CAS  Google Scholar 

  • Bayram O, Bayram OS, Ahmed YL, Maruyama J, Valerius O, Rizzoli SO, Ficner R, Irniger S, Braus GH (2012) The Aspergillus nidulans MAPK module AnSte11-Ste50-Ste7-Fus3 controls development and secondary metabolism. PLoS Genet 8:e1002816

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Bhattacharyya RP, Remenyi A, Good MC, Bashor CJ, Falick AM, Lim WA (2006) The Ste5 scaffold allosterically modulates signaling output of the yeast mating pathway. Science 311:822–826

    Article  PubMed  CAS  Google Scholar 

  • Bryksin AV, Matsumura I (2010) Overlap extension PCR cloning: a simple and reliable way to create recombinant plasmids. Biotechniques 48:463–465

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Cai Q, Wang JJ, Fu B, Ying SH, Feng MG (2018) Gcn5-dependent histone H3 acetylation and gene activity is required for the asexual development and virulence of Beauveria bassiana. Environ Microbiol 20:1484–1497

    Article  PubMed  CAS  Google Scholar 

  • Chen RE, Thorner J (2007) Function and regulation in MAPK signaling pathways: lessons learned from the yeast Saccharomyces cerevisia. Biochim Biophys Acta-Mol. Cell Res 1773:1311–1340

    CAS  Google Scholar 

  • Chen JY, Chen J, Lane S, Liu HP (2002) A conserved mitogen-activated protein kinase pathway is required for mating in Candida albicans. Mol Microbiol 46:1335–1344

    Article  PubMed  CAS  Google Scholar 

  • Davanture M, Dumur J, Bataillé-Simoneau N, Campion C, Valot B, Zivy M, Simoneau P, Fillinger S (2014) Phosphoproteome profiles of the phytopathogenic fungi Alternaria brassicicola and Botrytis cinerea during exponential growth in axenic cultures. Proteomics 14:1639–1645

    Article  PubMed  CAS  Google Scholar 

  • Davidson RC, Nicholls CB, Cox GM, Perfect JR, Heitman J (2003) A MAP kinase cascade composed of cell type specific and non-specific elements controls mating and differentiation of the fungal pathogen Cryptococcus neoformans. Mol Microbiol 49:469–485

    Article  PubMed  CAS  Google Scholar 

  • Errede B, Ge QY (1996) Feedback regulation of map kinase signal pathways. Philos Trans R Soc Lond Ser B Biol Sci 351:143–148

    Article  CAS  Google Scholar 

  • Etxebeste O, Garzia A, Espeso EA, Ugalde U (2010) Aspergillus nidulans asexual development: making the most of cellular modules. Trends Microbiol 18:569–576

    Article  PubMed  CAS  Google Scholar 

  • Franck WL, Gokce E, Randall SM, Oh Y, Eyre A, Muddiman DC, Dean RA (2015) Phosphoproteome analysis links protein phosphorylation to cellular remodeling and metabolic adaptation during Magnaporthe oryzae appressorium development. J Proteome Res 14:2408–2424

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Good M, Tang G, Singleton J, Remenyi A, Lim WA (2009) The Ste5 scaffold directs mating signaling by catalytically unlocking the Fus3 MAP kinase for activation. Cell 136:1085–1097

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Gu Q, Chen Y, Liu Y, Zhang CQ, Ma ZH (2015) The transmembrane protein FgSho1 regulates fungal development and pathogenicity via the MAPK module Ste50-Ste11-Ste7 in Fusarium graminearum. New Phytol 206:315–328

    Article  PubMed  CAS  Google Scholar 

  • Gustin MC, Albertyn J, Alexander M, Davenport K (1998) MAP kinase pathways in the yeast Saccharomyces cerevisiae. Microbiol Mol Biol Rev 62:1264–1300

    PubMed  PubMed Central  CAS  Google Scholar 

  • Hansen KC, Schmitt-Ulms G, Chalkley RJ, Hirsch J, Baldwin MA, Burlingame AL (2003) Mass spectrometric analysis of protein mixtures at low levels using cleavable C-13-isotope-coded affinity tag and multidimensional chromatography. Mol Cell Proteomics 2:299–314

    Article  PubMed  CAS  Google Scholar 

  • Holder DJ, Keyhani NO (2005) Adhesion of the entomopathogenic fungus Beauveria (Cordyceps) bassiana to substrata. Appl Environ Microbiol 71:5260–5266

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Huang BF, Feng MG (2009) Comparative tolerances of various Beauveria bassiana isolates to UV-B irradiation with a description of a modeling method to assess lethal dose. Mycopathologia 168:145–152

    Article  PubMed  Google Scholar 

  • Kim YK, Kawano T, Li DX, Kolattukudy PE (2000) A mitogen-activated protein kinase kinase required for induction of cytokinesis and appressorium formation by host signals in the conidia of Colletotrichum gloeosporioides. Plant Cell 12:1331–1343

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Kitade Y, Sumita T, Izumitsu K, Tanaka C (2015) MAPKK-encoding gene Ste7 in Bipolaris maydis is required for development and morphogenesis. Mycoscience 56:150–158

    Article  CAS  Google Scholar 

  • Li YF, Ye ZJ, Nie YF, Zhang J, Wang GL, Wang ZZ (2015a) Comparative phosphoproteome analysis of Magnaporthe oryzae-responsive proteins in susceptible and resistant rice cultivars. J Proteome 115:66–80

    Article  CAS  Google Scholar 

  • Li F, Shi HQ, Ying SH, Feng MG (2015b) WetA and VosA are distinct regulators of conidiation capacity, conidial quality, and biological control potential of a fungal insect pathogen. Appl Microbiol Biotechnol 99:10069–10081

    Article  PubMed  CAS  Google Scholar 

  • Li F, Shi HQ, Ying SH, Feng MG (2015c) Distinct contributions of one Fe- and two Cu/Zn-cofactored superoxide dismutases to antioxidation, UV tolerance and virulence of Beauveria bassiana. Fungal Genet Biol 81:160–171

    Article  PubMed  CAS  Google Scholar 

  • Liñeiro E, Chiva C, Cantoral JM, Sabido E, Fernández-Acero FJ (2016) Phosphoproteome analysis of B. cinerea in response to different plant-based elicitors. J Proteome 139:84–94

    Article  CAS  Google Scholar 

  • Liu J, Wang ZK, Sun HH, Ying SH, Feng MG (2017) Characterization of the Hog1 MAPK pathway in the entomopathogenic fungus Beauveria bassiana. Environ Microbiol 19:1808–1821

    Article  PubMed  CAS  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCt method. Methods 25:402–408

    Article  PubMed  CAS  Google Scholar 

  • Luo ZB, Qin YQ, Pei Y, Keyhani NO (2014) Ablation of the creA regulator results in amino acid toxicity, temperature sensitivity, pleiotropic effects on cellular development and loss of virulence in the filamentous fungus Beauveria bassiana. Environ Microbiol 16:1122–1136

    Article  PubMed  CAS  Google Scholar 

  • Ma QS, Jin K, Peng GX, Xia YX (2015) An ENA ATPase, MaENA1, of Metarhizium acridum influences the Na+-, thermo- and UV-tolerances of conidia and is involved in multiple mechanisms of stress tolerance. Fungal Genet Biol 83:68–77

    Article  PubMed  CAS  Google Scholar 

  • Maleri S, Ge Q, Hackett EA, Wang Y, Dohlman HG, Errede B (2004) Persistent activation by constitutive Ste7 promotes Kss1-mediated invasive growth but fails to support Fus3-dependent mating in yeast. Mol Cell Biol 24:9221–9238

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Martin H, Flandez M, Nombela C, Molina M (2005) Protein phosphatases in MAPK signalling: we keep learning from yeast. Mol Microbiol 58:6–16

    Article  PubMed  CAS  Google Scholar 

  • Müller P, Weinzierl G, Brachmann A, Feldbrügge M, Kahmann R (2003) Mating and pathogenic development of the Smut fungus Ustilago maydis are regulated by one mitogen-activated protein kinase cascade. Eukaryot Cell 2:1187–1199

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Park HS, Yu JH (2012) Genetic control of asexual sporulation in filamentous fungi. Curr Opin Microbiol 15:669–677

    Article  PubMed  CAS  Google Scholar 

  • Rampitsch C, Subramaniam R, Djuric-Ciganovic S, Bykova NV (2010) The phosphoproteome of Fusarium graminearum at the onset of nitrogen starvation. Proteomics 10:124–140

    Article  PubMed  CAS  Google Scholar 

  • Rampitsch C, Tinker NA, Subramaniam R, Barkow-Oesterreicher S, Laczko E (2012) Phosphoproteome profile of Fusarium graminearum grown in vitro under nonlimiting conditions. Proteomics 12:1002–1005

    Article  PubMed  CAS  Google Scholar 

  • Ramsubramaniam N, Harris SD, Marten MR (2014) The phosphoproteome of Aspergillus nidulans reveals functional association with cellular processes involved in morphology and secretion. Proteomics 14:2454–2459

    Article  PubMed  CAS  Google Scholar 

  • Rispail N, Soanes DM, Ant C, Czajkowski R, Grünler A, Huguet R, Perez-Nadales E, Poli A, Sartorel E, Valiante V, Yang M, Beffa R, Brakhage AA, Gow NAR, Kahmann R, Lebrun MH, Lenasi H, Perez-Martin J, Talbot NJ, Wendland J, Di Pietro A (2009) Comparative genomics of MAP kinase and calcium-calcineurin signalling components in plant and human pathogenic fungi. Fungal Genet Biol 46:287–298

    Article  PubMed  CAS  Google Scholar 

  • Rodriguez-Navarro A, Benito B (2010) Sodium or potassium efflux ATPase: a fungal, bryophyte, and protozoal ATPase BBA. Biomembranes 1798:1841–1853

    Article  CAS  Google Scholar 

  • Sakaguchi A, Tsuji G, Kubo Y (2010) A yeast STE11 homologue CoMEKK1 is essential for pathogenesis-related morphogenesis in Colletotrichum orbiculare. Mol Plant-Microbe Interact 23:1563–1572

    Article  PubMed  CAS  Google Scholar 

  • Schamber A, Leroch M, Diwo J, Mendgen K, Hahn M (2010) The role of mitogen-activated protein (MAP) kinase signalling components and the Ste12 transcription factor in germination and pathogenicity of Botrytis cinerea. Mol Plant Pathol 11:105–119

    Article  PubMed  CAS  Google Scholar 

  • Tong SM, Zhang AX, Guo CT, Ying SH, Feng MG (2018) Daylight length-dependent translocation of VIVID photoreceptor in cells and its essential role in conidiation and virulence of Beauveria bassiana. Environ Microbiol 20:169–185

    Article  PubMed  CAS  Google Scholar 

  • van Drogen F, Stucke VM, Jorritsma G, Peter M (2001) MAP kinase dynamics in response to pheromones in budding yeast. Nat Cell Biol 3:1051–1059

    Article  PubMed  CAS  Google Scholar 

  • Wang J, Liu J, Hu Y, Ying SH, Feng MG (2013a) Cytokinesis-required Cdc14 is a signaling hub of asexual development and multi-stress tolerance in Beauveria bassiana. Sci Rep 3:3086

    Article  PubMed  PubMed Central  Google Scholar 

  • Wang ZL, Zhang LB, Ying SH, Feng MG (2013b) Catalases play differentiated roles in the adaptation of a fungal entomopathogen to environmental stresses. Environ Microbiol 15:409–418

    Article  PubMed  CAS  Google Scholar 

  • Wang ZK, Wang J, Liu J, Ying SH, Peng XJ, Feng MG (2016a) Proteomic and phosphoproteomic insights into a signaling hub role for Cdc14 in asexual development and multiple stress responses in Beauveria bassiana. PLoS One 11:e0153007

    Article  PubMed  PubMed Central  Google Scholar 

  • Wang J, Ying SH, Hu Y, Feng MG (2016b) Mas5, a homologue of bacterial DnaJ, is indispensable for the host infection and environmental adaptation of a filamentous fungal insect pathogen. Environ Microbiol 18:1037–1047

    Article  PubMed  CAS  Google Scholar 

  • Xiao GH, Ying SH, Zheng P, Wang ZL, Zhang SW, Xie XQ, Shang YF, St Leger RJ, Zhao GP, Wang CS, Feng MG (2012) Genomic perspectives on the evolution of fungal entomopathogenicity in Beauveria bassiana. Sci Rep 2:483

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Xie XQ, Li F, Ying SH, Feng MG (2012) Additive contributions of two manganese-cored superoxide dismutases (MnSODs) to antioxidation, UV tolerance and virulence of Beauveria bassiana. PLoS One 7:e30298

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Xiong Y, Coradetti ST, Li X, Gritsenko MA, Clauss T, Petyuk V, Camp D, Smith R, Cate JHD, Yang F, Glass NL (2014) The proteome and phosphoproteome of Neurospora crassa in response to cellulose, sucrose and carbon starvation. Fungal Genet Biol 72:21–33

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Ying SH, Feng MG (2006) Novel blastospore-based transformation system for integration of phosphinothricin resistance and green fluorescence protein genes into Beauveria bassiana. Appl Microbiol Biotechnol 72:206–210

    Article  PubMed  CAS  Google Scholar 

  • Ying SH, Feng MG, Keyhani NO (2013) Use of uridine auxotrophy (ura3) for markerless transformation of the mycoinsecticide Beauveria bassiana. Appl Microbiol Biotechnol 97:3017–3025

    Article  PubMed  CAS  Google Scholar 

  • Zhang LB, Feng MG (2018) Antioxidant enzymes and their contributions to biological control potential of fungal insect pathogens. Appl Microbiol Biotechnol 102:4995–5004

    Article  PubMed  CAS  Google Scholar 

  • Zhang YJ, Zhang JQ, Jiang XD, Wang GJ, Luo ZB, Fan YH, Wu ZQ, Pei Y (2010) Requirement of a mitogen-activated protein kinase for appressorium formation and penetration of insect cuticle by the entomopathogenic fungus Beauveria bassiana. Appl Environ Microbiol 76:2262–2270

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Zhao X, Kim Y, Park G, Xu JR (2005) A mitogen-activated protein kinase cascade regulating infection-related morphogenesis in Magnaporthe grisea. Plant Cell 17:1317–1329

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Zheng CF, Guan KL (1994) Activation of MEK family kinases requires phosphorylation of two conserved Ser/Thr residues. EMBO J 13:1123–1131

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Zhu J, Ying SH, Feng MG (2016) The Na+/H+ antiporter Nhx1 controls vacuolar fusion indispensible for the life cycle in vitro and in vivo of a fungal insect pathogen. Environ Microbiol 18:3884–3895

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Natural Science Foundation of China (Grant No. 31572054), the Ministry of Science and Technology of the People’s Republic of China (Grant No. 2017YFD0201202), and the Fundamental Research Funds for the Central Universities (Grant No. 2018FZA6003).

Author information

Author notes

  1. Zhi-Kang Wang and Qing Cai contributed equally to this work.

Authors and Affiliations

  1. Institute of Coastal Ecology, Ludong University, Yantai, 264025, Shandong, China

    Zhi-Kang Wang

  2. Institute of Microbiology, College of Life Sciences, Zhejiang University, Zhejiang, 310058, Hangzhou, China

    Zhi-Kang Wang, Qing Cai, Sen-Miao Tong, Sheng-Hua Ying & Ming-Guang Feng

  3. School of Agricultural and Food Science, Zhejiang A&F University, Lin’an, Zhejiang, 311300, China

    Sen-Miao Tong

Authors
  1. Zhi-Kang Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Qing Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sen-Miao Tong
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Sheng-Hua Ying
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ming-Guang Feng
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ming-Guang Feng.

Ethics declarations

Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Electronic supplementary material

ESM 1

(PDF 4295 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, ZK., Cai, Q., Tong, SM. et al. C-terminal Ser/Thr residues are vital for the regulatory role of Ste7 in the asexual cycle and virulence of Beauveria bassiana. Appl Microbiol Biotechnol 102, 6973–6986 (2018). https://doi.org/10.1007/s00253-018-9148-5

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-018-9148-5

Keywords

Search

Navigation