The protein phosphatase gene MaPpt1 acts as a programmer of microcycle conidiation and a negative regulator of UV-B tolerance in Metarhizium acridum

  • Jie Zhang
  • Zhenglong Wang
  • Nemat O. Keyhani
  • Guoxiong Peng
  • Kai JinEmail author
  • Yuxian XiaEmail author
Applied genetics and molecular biotechnology


The Ser/Thr protein phosphatase Ppt1 (yeast)/PP5 (humans) has been implicated in signal transduction–mediated growth and differentiation, DNA damage/repair, cell cycle progression, and heat shock responses. Little, however, is known concerning the functions of Ppt1/PP5 in filamentous fungi. In this study, the Ppt1 gene MaPpt1 was characterized in the insect pathogenic fungus, Metarhizium acridum. The MaPpt1 protein features a three-tandem tetratricopeptide repeat (TPR) domain and a peptidyl-prolyl cis-trans isomerase-like (PP2Ac) domain. Subcellular localization using an MaPpt1::eGFP fusion protein revealed that MaPpt1 was localized in the cytoplasm of spores, but gathered at the septa in growing hyphae. Targeted gene inactivation of MaPpt1 in M. acridum resulted in unexpected reprogramming of normal aerial conidiation to microcycle conidiation. Although overall vegetative growth was unaffected, a significant increase in conidial yield was noted in ΔMaPpt1. Stress-responsive phenotypes and virulence were largely unaffected in ΔMaPpt1. Exceptionally, ΔMaPpt1 displayed increased UV tolerance compared to wild type. Digital gene expression data revealed that MaPpt1 mediates transcription of sets of genes involved in conidiation, polarized growth, cell cycle, cell proliferation, DNA replication and repair, and some important signaling pathways. These data indicate a unique role for Ppt1 in filamentous fungal development and differentiation.


Protein phosphatase Ppt1 Metarhizium acridum Microcycle conidiation UV tolerance 



This study was funded by Natural Science Foundation of China (No. 31272090), the Natural Science Foundation Project of Chongqing (CSTC 2018jcyjAX0554), and the Fundamental Research Funds for the Central Universities (106112017CDJQJ298831).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

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

Supplementary material

253_2018_9567_MOESM1_ESM.pdf (1.4 mb)
ESM 1 (PDF 1386 kb)


  1. Ali A, Zhang J, Bao S, Liu I, Otterness D, Dean NM, Abraham RT, Wang XF (2004) Requirement of protein phosphatase 5 in DNA-damage-induced ATM activation. Genes Dev 18:249–254CrossRefGoogle Scholar
  2. Audic S, Claverie JM (1997) The significance of digital gene expression profiles. Genome Res 7:986–995CrossRefGoogle Scholar
  3. Azam M, Kesarwani M, Natarajan K, Datta A (2001) A secretion signal is present in the Collybia velutipes oxalate decarboxylase gene. Biochem Biophys Res Commun 289:807–812CrossRefGoogle Scholar
  4. Barford D (1996) Molecular mechanisms of the protein Ser/Thr phosphatases. Trends Biochem Sci 21:407–412CrossRefGoogle Scholar
  5. Becker W, Kentrup H, Klumpp S, Schultz JE, Joost HG (1994) Molecular cloning of a protein serine/threonine phosphatase containing a putative regulatory tetratricopeptide repeat domain. J Biol Chem 269:22586–22592PubMedGoogle Scholar
  6. Bosch A, Yantorno O (1999) Microcycle conidiation in the entomopathogenic fungus Beauveria bassiana bals. (Vuill.). Process Biochem 34:707–716CrossRefGoogle Scholar
  7. Chaudhuri M (2001) Cloning and characterization of a novel serine/threonine protein phosphatase type 5 from Trypanosoma brucei. Gene 266:1–13CrossRefGoogle Scholar
  8. Chen MX, McPartlin AE, Brown L, Chen YH, Barker HM, Cohen P (1994) A novel human protein serine/threonine phosphatase, which possesses four tetratricopeptide repeat motifs and localizes to the nucleus. EMBO J 13:4278–4290CrossRefGoogle Scholar
  9. Chinkers M (2001) Protein phosphatase 5 in signal transduction. Trends Endocrinol Metab 12:28–32CrossRefGoogle Scholar
  10. D’Errico M, Parlanti E, Pascucci B, Fortini P, Baccarini S, Simonelli V, Dogliotti E (2017) Single nucleotide polymorphisms in DNA glycosylases: from function to disease. Free Radic Biol Med 107:278–291CrossRefGoogle Scholar
  11. Das AK, Cohen PTW, Barford D (1998) The structure of the tetratricopeptide repeats of protein phosphatase 5: implications for TPR-mediated protein–protein interactions. EMBO J 17:1192–1199CrossRefGoogle Scholar
  12. Dickman MB, Yarden O (1999) Serine/threonine protein kinases and phosphatases in filamentious fungi. Fungal Genet Biol 26:99–117CrossRefGoogle Scholar
  13. Diernfellner ACR, Schafmeier T (2011) Phosphorylations: making the Neurospora crassa circadian clock tick. FEBS Lett 585:1461–1466CrossRefGoogle Scholar
  14. Doherty AJ, Wigley DB (1999) Functional domains of an ATP-dependent DNA ligase. J Mol Biol 285:63–71CrossRefGoogle Scholar
  15. dos Reis MC, Pelegrinelli Fungaro MH, Delgado Duarte RT, Furlaneto L, Furlaneto MC (2004) Agrobacterium tumefaciens-mediated genetic transformation of the entomopathogenic fungus Beauveria bassiana. J Microbiol Methods 58:97–202Google Scholar
  16. Du Y, Jin K, Xia Y (2018) Involvement of MaSom1, a downstream transcriptional factor of cAMP/PKA pathway, in conidial yield, stress tolerances, and virulence in Metarhizium acridum. Appl Microbiol Biotechnol 102:5611–5623CrossRefGoogle Scholar
  17. Eberle RJ, Coronado MA, Caruso IP, Lopes DO, Miyoshi A, Azevedo V, Arni RK (2015) Chemical and thermal influence of the [4Fe–4S]2+ cluster of A/G-specific adenine glycosylase from Corynebacterium pseudotuberculosis. Biochim Biophys Acta 1850:393–400CrossRefGoogle Scholar
  18. Emptage K, O'Neill R, Solovyova A, Connolly BA (2008) Interplay between DNA polymerase and proliferating cell nuclear antigen switches off base excision repair of uracil and hypoxanthine during replication in archaea. J Mol Biol 383:762–771CrossRefGoogle Scholar
  19. Feng B, Zhao C, Tanaka S, Imanaka HK, Nakanishi K (2007) TPR domain of Ser/Thr phosphatase of Aspergillus oryzae shows no auto-inhibitory effect on the dephosphorylation activity. Int J Biol Macromol 41:281–285CrossRefGoogle Scholar
  20. Fumasoni M, Zwicky K, Vanoli F, Lopes M, Branzei D (2015) Error-free DNA damage tolerance and sister chromatid proximity during DNA replication rely on the Polα/Primase/Ctf4 Complex. Mol Cell 57:812–823CrossRefGoogle Scholar
  21. Gao Q, Jin K, Ying S, Zhang Y, Xiao G, Shang Y, Duan Z, Hu X, Xie X, Zhou G, Peng G, Luo Z, Huang W, Wang B, Fang W, Wang S, Zhong Y, Ma L, St Leger RJ, Zhao G, Pei Y, Feng M, Xia Y, Wang C (2011) Genome sequencing and comparative transcriptomics of the model entomopathogenic fungi Metarhizium anisopliae and M. acridum. PLoS Genet 7:e1001264CrossRefGoogle Scholar
  22. Geymonat M, Spanos A, Jensen S, Sedgwick SG (2010) Phosphorylation of Lte1 by Cdk prevents polarized growth during mitotic arrest in S. cerevisiae. J Cell Biol 191:1097–1112Google Scholar
  23. Guo JH, Cheng P, Liu Y (2010) Functional significance of FRH in regulating the phosphorylation and stability of Neurospora circadian clock protein FRQ. J Biol Chem 285:11508–11,515CrossRefGoogle Scholar
  24. Hašplová K, Hudecová A, Magdolénová Z, Bjøras M, Gálová E, Miadoková E, Dušinská M (2012) DNA alkylation lesions and their repair in human cells: modification of the comet assay with 3-methyladenine DNA glycosylase (AlkD). Toxicol Lett 208:76–81CrossRefGoogle Scholar
  25. Hu K, Li W, Gao J, Liu Q, Wang H, Wang Y, Sang J (2014) Role of Ppt1 in multiple stress responses in Candida albicans. Chin Sci Bull 59:4060–4068CrossRefGoogle Scholar
  26. Jeong JY, Johns J, Sinclair C, Park JM, Rossie S (2003) Characterization of Saccharomyces cerevisiae protein Ser/Thr phosphatase T1 and comparison to its mammalian homolog PP5. BMC Cell Biol 4(3):3CrossRefGoogle Scholar
  27. Kang Y, Cheong HM, Lee JH, Song PI, Lee KH, Kim SY, Jun JY, You HJ (2011) Protein phosphatase 5 is necessary for ATR-mediated DNA repair. Biochem Biophys Res Commun 404:476–481CrossRefGoogle Scholar
  28. Kennelly PJ (2001) Protein phosphatases--a phylogenetic perspective. Chem Rev 101:2291–2312CrossRefGoogle Scholar
  29. de la Fuente van Bentem S, Vossen JH, Vermeer JE, de Vroomen MJ, Gadella TW, Haring MA, Cornelissen BJ (2003) The subcellular localization of plant protein phosphatase 5 isoforms is determined by alternative splicing. Plant Physiol 133:702–712CrossRefGoogle Scholar
  30. Lazo GR, Stein PA, Ludwig RA (1991) A DNA transformation competent Arabidopsis genomic library in Agrobacterium. Biotechnology 9:963–967CrossRefGoogle Scholar
  31. Lee AJ, Wallace SS (2016) Visualizing the search for radiation-damaged DNA bases in real time. Radiat Phys Chem Oxf Engl 1993 128:126–133PubMedPubMedCentralGoogle Scholar
  32. Li ZZ, Alves SB, Roberts DW, Fan MZ, Delalibera I, Tang J, Lopes RB, Faria M, Rangel DEN (2010) Biological control of insects in Brazil and China: history, current programs and reasons for their successes using entomopathogenic fungi. Biocontrol Sci Tech 20:117–136CrossRefGoogle Scholar
  33. Liu J, Cao Y, Xia Y (2010) Mmc, a gene involved in microcycle conidiation of the entomopathogenic fungus Metarhizium anisopliae. J Invertebr Pathol 105:132–138CrossRefGoogle Scholar
  34. Liu X, Li H, Liu Q, Niu Y, Hu Q, Deng H, Cha J, Wang Y, Liu Y, He Q (2015) Role for protein kinase A in the Neurospora circadian clock by regulating white collar-independent frequency transcription through phosphorylation of RCM-1. Mol Cell Biol 35:2088–2102CrossRefGoogle Scholar
  35. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCTmethod. Methods 25:402–408PubMedPubMedCentralGoogle Scholar
  36. Lomer CJ, Bateman RP, Johnson DL, Langewald J, Thomas M (2001) Biological control of locusts and grasshoppers. Annu Rev Entomol 46:667–702CrossRefGoogle Scholar
  37. Luo S, He M, Cao Y, Xia Y (2013) The tetraspanin gene MaPls1 contributes to virulence by affecting germination, appressorial function and enzymes for cuticle degradation in the entomopathogenic fungus Metarhizium acridum. Environ Microbiol 15:2966–2979PubMedGoogle Scholar
  38. Ma GX, Rong QZ, Shi JH, Han CH, Tao Z, Xia QY (2014) Molecular characterization and functional analysis of serine/threonine protein phosphatase of Toxocara canis. Exp Parasitol 141:55–61CrossRefGoogle Scholar
  39. Maheshwari R (1991) Microcycle conidiation and its genetic basis in Neurospora crassa. J Gen Microbiol 137:2103–2115CrossRefGoogle Scholar
  40. Ming Y, Wei Q, Jin K, Xia Y (2014) MaSnf1, a sucrose non-fermenting protein kinase gene, is involved in carbon source utilization, stress tolerance, and virulence in Metarhizium acridum. Appl Microbiol Biotechnol 98:10153–10164CrossRefGoogle Scholar
  41. Monje-Casas F, Amon A (2009) Cell polarity determinants establish asymmetry in MEN signaling. Dev Cell 16:132–145CrossRefGoogle Scholar
  42. Ortiz-Urquiza A, Keyhani NO (2013) Action on the surface: entomopathogenic fungi versus the insect cuticle. Insects 4:357–374CrossRefGoogle Scholar
  43. Ortiz-Urquiza A, Luo Z, Keyhani NO (2015) Improving mycoinsecticides for insect biological control. Appl Microbiol Biotechnol 99:1057–1068CrossRefGoogle Scholar
  44. Rangel DEN, Braga GUL, Fernandes ÉKK, Keyser CA, Hallsworth JE, Roberts DW (2015) Stress tolerance and virulence of insect-pathogenic fungi are determined by environmental conditions during conidial formation. Curr Genet 61:383–404CrossRefGoogle Scholar
  45. Rödel C, Jupitz T, Schmidt H (1999) Complementation of the DNA repair-deficient swi10 mutant of fission yeast by the human ERCC1 gene. Nucleic Acids Res 25:2823–2827CrossRefGoogle Scholar
  46. Rodriguez-Urra AB, Jimenez C, Nieto MI, Rodriguez J, Hayashi H, Ugalde U (2012) Signaling the induction of sporulation involves the interaction of two secondary metabolites in Aspergillus nidulans. ACS Chem Biol 7:599–606CrossRefGoogle Scholar
  47. Schmoll M, Dattenböck C, Carrerasvillaseñor N , Mendozamendoza A , Tisch D, Alemán M I, , Baker SE, Brown C, Cervantes-Badillo MG, Cetz-Chel J, Cristobal-Mondragon GR, Delaye L, Esquivel-Naranjo EU, Frischmann A, Gallardo-Negrete Jde J, García-Esquivel M, Gomez-Rodriguez EY, Greenwood DR, Hernández-Oñate M, Kruszewska JS, Lawry R, Mora-Montes HM, Muñoz-Centeno T, Nieto-Jacobo MF, Nogueira Lopez G, Olmedo-Monfil V, Osorio-Concepcion M, Piłsyk S, Pomraning KR, Rodriguez-Iglesias A, Rosales-Saavedra MT, Sánchez-Arreguín JA, Seidl-Seiboth V, Stewart A, Uresti-Rivera EE, Wang CL, Wang TF, Zeilinger S, Casas-Flores S, Herrera-Estrella A (2016) The genomes of three uneven siblings: footprints of the lifestyles of three Trichoderma species. Microbiol Mol Biol Rev 80:205–327CrossRefGoogle Scholar
  48. Schroeder A (1975) Genetic control of radiation sensitivity and DNA repair in Neurospora. Basic Life Sci 5B:567–576PubMedGoogle Scholar
  49. Schwede T, Kopp J, Guex N, Peitsch M (2003) SWISS-MODEL: an automated protein homology-modeling server. Nucleic Acids Res 31:3381–3385CrossRefGoogle Scholar
  50. Seshan A, Bardin AJ, Amon A (2002) Control of Lte1 localization by cell polarity determinants and Cdc14. Curr Biol 12:2098–2110CrossRefGoogle Scholar
  51. Shi Y (2009) Serine/threonine phosphatases: mechanism through structure. Cell 139:468–484CrossRefGoogle Scholar
  52. Sinha RPS, Häder DP (2002) UV-induced DNA damage and repair: a review. Photochem Photobiol Sci 1:225–236CrossRefGoogle Scholar
  53. Smits GJ, Schenkman LR, Brul S, Pringle JR, Klis FM (2006) Role of cell cycle-regulated expression in the localized incorporation of cell wall proteins in yeast. Mol Biol Cell 17:3267–3280CrossRefGoogle Scholar
  54. Troll CJ, Adhikary S, Cueff M, Mitra I, Eichman BF, Camps M (2014) Interplay between base excision repair activity and toxicity of 3-methyladenine DNA glycosylases in an E. coli complementation system. Mutat Res 763–764: 64–73, 763-764Google Scholar
  55. Venkannagari H, Verheugd P, Koivunen J, Haikarainen T, Obaji E, Ashok Y, Narwal M, Pihlajaniemi T, Lüscher B, Lehtiö L (2016) Small-molecule chemical probe rescues cells from mono-ADP-ribosyltransferase ARTD10/PARP10-induced apoptosis and sensitizes cancer cells to DNA damage. Cell Chem Biol 23:1251–1260CrossRefGoogle Scholar
  56. Wandinger SK, Suhre MH, Wegele H, Buchner J (2006) The phosphatase Ppt1 is a dedicated regulator of the molecular chaperone Hsp90. EMBO J 25:367–376CrossRefGoogle Scholar
  57. Wei Q, Li JH, Liu T, Tong XM, Ye X (2013) Phosphorylation of minichromosome maintenance protein 7 (MCM7) by cyclin/cyclin-dependent kinase affects its function in cell cycle regulation. J Biol Chem 288:19715–19725CrossRefGoogle Scholar
  58. Yong W, Bao S, Chen H, Li D, Sánchez ER, Shou W (2007) Mice lacking protein phosphatase 5 are defective in ataxia telangiectasia mutated (ATM)-mediated cell cycle arrest. J Biol Chem 282:14690–14694CrossRefGoogle Scholar
  59. Yoshida K, Inoue I (2003) Conditional expression of MCM7 increases tumor growth without altering DNA replication activity. FEBS Lett 553:213–217CrossRefGoogle Scholar
  60. Zhang LB, Feng MG (2018) Antioxidant enzymes and their contributions to biological control potential of fungal insect pathogens. Appl Microbiol Biotechnol 102:4995–5004CrossRefGoogle Scholar
  61. Zhang J, Bao S, Furumai R, Kucera KS, Ali A, Dean NM, Wang XF (2005) Protein phosphatase 5 is required for ATR-mediated checkpoint activation. Mol Cell Biol 25:9910–9919CrossRefGoogle Scholar
  62. Zhang S, Peng G, Xia Y (2010) Microcycle conidiation and the conidial properties in the entomopathogenic fungus Metarhizium acridum on agar medium. Biocontrol Sci Tech 20:809–819CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Genetic Engineering Research Center, School of Life SciencesChongqing UniversityChongqingPeople’s Republic of China
  2. 2.Chongqing Engineering Research Center for Fungal InsecticideChongqingPeople’s Republic of China
  3. 3.Key Laboratory of Gene Function and Regulation Technologies under Chongqing Municipal Education CommissionChongqingPeople’s Republic of China
  4. 4.Department of Microbiology and Cell ScienceUniversity of FloridaGainesvilleUSA

Personalised recommendations