Skip to main content
SpringerLink
Log in

The NcZrg-17 gene of Neurospora crassa encodes a cation diffusion facilitator transporter required for vegetative development, tolerance to endoplasmic reticulum stress and cellulose degradation under low zinc conditions

  • Original Article
  • Published:
Current Genetics Aims and scope Submit manuscript

Abstract

The Neurospora crassa gene NcZrg-17 encodes a membrane protein with homology to the cation diffusion facilitator (CDF) family of transporters. We analyzed the phenotypic and functional characteristics of ΔNcZrg-17 and the implications of these characteristics in vivo. The ΔNcZrg-17 mutant showed several phenotypes that are zinc suppressible such as reduced growth rate, short aerial hyphae, increased hyphal branching, early and enhanced conidiation and delayed conidial germination. Furthermore, the NcZrg-17 gene was found to be crucial for survival in the presence of endoplasmic reticulum (ER) stress inducing chemical agents. In addition, we found that ΔNcZrg-17 mutant is defective in protein secretion on cellulose media under low zinc conditions, pointing towards a physiological role for NcZrg-17 in N. crassa. A gradual and delayed transcriptional upregulation (~ threefold) of NcZrg-17 on exposure to low zinc suggests its role in adaptation to low zinc rather than zinc homeostasis. Together our findings support a function of NcZrg-17 in normal vegetative growth, tolerance to ER stress and degradation of cellulose under low zinc conditions in N. crassa.

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

  • Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410

    Article  PubMed  CAS  Google Scholar 

  • Arias P, Díez-Muñiz S, García R, Nombela C, Rodríguez-Peña JM, Arroyo J (2011) Genome-wide survey of yeast mutations leading to activation of the yeast cell integrity MAPK pathway: novel insights into diverse MAPK outcomes. BMC Genom 12:1

    Article  CAS  Google Scholar 

  • Barman A, Tamuli R (2015) Multiple cellular roles of Neurospora crassa plc-1, splA2, and cpe-1 in regulation of cytosolic free calcium, carotenoid accumulation, stress responses, and acquisition of thermotolerance. J Microbiol 53:226–235

    Article  PubMed  CAS  Google Scholar 

  • Benz JP, Chau BH, Zheng D, Bauer S, Glass NL, Somerville CR (2014) A comparative systems analysis of polysaccharide-elicited responses in Neurospora crassa reveals carbon source-specific cellular adaptations. Mol Microbiol 91:275–299

    Article  PubMed  CAS  Google Scholar 

  • Beseli A, Amnuaykanjanasin A, Herrero S, Thomas E, Daub ME (2015) Membrane transporters in self resistance of Cercospora nicotianae to the photoactivated toxin cercosporin. Curr Genet 61:601–620

    Article  PubMed  CAS  Google Scholar 

  • Colot HV, Park G, Turner GE, Ringelberg C, Crew CM, Litvinkova L, Weiss RL, Borkovich KA, Dunlap JC (2006) A high-throughput gene knockout procedure for Neurospora reveals functions for multiple transcription factors. Proc Natl Acad Sci 103:10352–10357

    Article  PubMed  CAS  Google Scholar 

  • Davis RH, de Serres FJ (1970) Genetic and microbiological research techniques for Neurospora crassa. Methods Enzymol 17:79–143

    Article  Google Scholar 

  • Deka R, Kumar R, Tamuli R (2011) Neurospora crassa homologue of Neuronal Calcium Sensor-1 has a role in growth, calcium stress tolerance, and ultraviolet survival. Genetica 139(7):885–894

    Article  PubMed  CAS  Google Scholar 

  • Eide D, Broderius M, Fett J, Guerinot ML (1996) A novel iron-regulated metal transporter from plants identified by functional expression in yeast. Proc Natl Acad Sci 93:5624–5628

    Article  PubMed  CAS  Google Scholar 

  • Ellis CD, Wang F, MacDiarmid CW, Clark S, Lyons T, Eide DJ (2004) Zinc and the Msc2 zinc transporter protein are required for endoplasmic reticulum function. J Cell Biol 166:325–335

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Ellis CD, MacDiarmid CW, Eide DJ (2005) Heteromeric protein complexes mediate zinc transport into the secretory pathway of eukaryotic cells. J Biol Chem 280:28811–28818

    Article  PubMed  CAS  Google Scholar 

  • Fan F, Ma G, Li J, Liu Q, Benz JP, Tian C, Ma Y (2015) Genome-wide analysis of the endoplasmic reticulum stress response during lignocellulase production in Neurospora crassa. Biotechnol Biofuels 8:1

    Article  CAS  Google Scholar 

  • Fang Y, Sugiura R, Ma Y, Yada-Matsushima T, Umeno H, Kuno T (2008) Cation diffusion facilitator Cis4 is implicated in Golgi membrane trafficking via regulating zinc homeostasis in fission yeast. Mol Biol Cell 19:1295–1303

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Fukada T, Kambe T (2011) Molecular and genetic features of zinc transporters in physiology and pathogenesis. Metallomics 3:662–674

    Article  PubMed  CAS  Google Scholar 

  • Guerinot ML (2000) The ZIP family of metal transporters. Biochim Biophys Acta (BBA) Biomembr 1465:190–198

    Article  CAS  Google Scholar 

  • Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. In: Nucleic acids symposium series, pp 95–98

  • Hill TW, Loprete DM, Momany M, Ha Y, Harsch LM, Livesay JA, Mirchandani A, Murdock JJ, Vaughan MJ, Watt MB (2006) Isolation of cell wall mutants in Aspergillus nidulans by screening for hypersensitivity to Calcofluor White. Mycologia 98:399–409

    Article  PubMed  CAS  Google Scholar 

  • Ishihara K, Yamazaki T, Ishida Y, Suzuki T, Oda K, Nagao M, Yamaguchi-Iwai Y, Kambe T (2006) Zinc transport complexes contribute to the homeostatic maintenance of secretory pathway function in vertebrate cells. J Biol Chem 281:17743–17750

    Article  PubMed  CAS  Google Scholar 

  • Kiranmayi P, Mohan PM (2006) Metal transportome of Neurospora crassa. In Silico Biol 6:169–180

    PubMed  CAS  Google Scholar 

  • Kiranmayi P, Tiwari A, Sagar KP, Haritha A, Maruthi Mohan P (2009) Functional characterization of tzn1 and tzn2-zinc transporter genes in Neurospora crassa. Biometals 22:411–420

    Article  PubMed  CAS  Google Scholar 

  • Korripally P, Tiwari A, Haritha A, Kiranmayi P, Bhanoori M (2010) Characterization of Ctr family genes and the elucidation of their role in the life cycle of Neurospora crassa. Fungal Genet Biol 47:237–245

    Article  PubMed  CAS  Google Scholar 

  • Kyte J, Doolittle RF (1982) A simple method for displaying the hydropathic character of a protein. J Mol Biol 157:105–132

    Article  PubMed  CAS  Google Scholar 

  • Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R (2007) Clustal W and clustal X version 2.0. Bioinformatics 23:2947–2948

    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. https://doi.org/10.1006/meth.2001.1262

  • MacDiarmid CW, Milanick MA, Eide DJ (2003) Induction of the ZRC1 metal tolerance gene in zinc-limited yeast confers resistance to zinc shock. J Biol Chem 278:15065–15072

    Article  PubMed  CAS  Google Scholar 

  • Malavazi I, Goldman GH, Brown NA (2014) The importance of connections between the cell wall integrity pathway and the unfolded protein response in filamentous fungi. Brief Funct Genom 13:456–470

    Article  CAS  Google Scholar 

  • Malhotra JD, Kaufman RJ (2007) Endoplasmic reticulum stress and oxidative stress: a vicious cycle or a double-edged sword? Antioxid Redox Signal 9:2277–2294

    Article  PubMed  CAS  Google Scholar 

  • Malhotra JD, Miao H, Zhang K, Wolfson A, Pennathur S, Pipe SW, Kaufman RJ (2008) Antioxidants reduce endoplasmic reticulum stress and improve protein secretion. Proc Natl Acad Sci 105:18525–18530

    Article  PubMed  Google Scholar 

  • McCluskey K, Wiest A, Plamann M (2010) The fungal genetics stock center: a repository for 50 years of fungal genetics research. J Biosci 35:119

    Article  PubMed  CAS  Google Scholar 

  • Mohan PM, Sastry KS (1983) Interrelationships in trace-element metabolism in metal toxicities in nickel-resistant strains of Neurospora crassa. Biochem J 212:205–210

    Article  Google Scholar 

  • Montanini B, Blaudez D, Jeandroz S, Sanders D, Chalot M (2007) Phylogenetic and functional analysis of the cation diffusion facilitator (CDF) family: improved signature and prediction of substrate specificity. BMC Genom 8:107

    Article  CAS  Google Scholar 

  • Montenegro-Montero A, Goity A, Larrondo LF (2015) The bZIP transcription factor HAC-1 is involved in the unfolded protein response and is necessary for growth on cellulose in Neurospora crassa. PLoS One 10:e0131415

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Nies DH (2003) Efflux-mediated heavy metal resistance in prokaryotes. FEMS Microbiol Rev 27:313–339

    Article  PubMed  CAS  Google Scholar 

  • Omasits U, Ahrens CH, Müller S, Wollscheid B (2013) Protter: interactive protein feature visualization and integration with experimental proteomic data. Bioinformatics 30:884–886

    Article  PubMed  CAS  Google Scholar 

  • Ouyang S, Beecher CN, Wang K, Larive CK, Borkovich KA (2015) Metabolic impacts of using nitrogen and copper-regulated promoters to regulate gene expression in Neurospora crassa. G3 Genes Genomes Genet 5:1899–1908

    CAS  Google Scholar 

  • Schneider CA, Rasband WS, Eliceiri KW (2012) NIH image to ImageJ: 25 years of image analysis. Nat Methods 9:671–675

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Suzuki T, Ishihara K, Migaki H, Ishihara K, Nagao M, Yamaguchi-Iwai Y, Kambe T (2005a) Two different zinc transport complexes of cation diffusion facilitator proteins localized in the secretory pathway operate to activate alkaline phosphatases in vertebrate cells. J Biol Chem 280:30956–30962

    Article  PubMed  CAS  Google Scholar 

  • Suzuki T, Ishihara K, Migaki H, Matsuura W, Kohda A, Okumura K, Nagao M, Yamaguchi-Iwai Y, Kambe T (2005b) Zinc transporters, ZnT5 and ZnT7, are required for the activation of alkaline phosphatases, zinc-requiring enzymes that are glycosylphosphatidylinositol-anchored to the cytoplasmic membrane. J Biol Chem 280:637–643

    Article  PubMed  CAS  Google Scholar 

  • RStudio Team (2015) RStudio: integrated Development for R. RStudio, Inc., Boston, MA. http://www.rstudio.com/

  • Tiwari A, Korripally P, Adhikarla H, Patnala K, Pamarthi MM, Bhanoori M (2011) Expression and functional characterisation of TNC, a high-affinity nickel transporter from Neurospora crassa. Fungal Genet Biol 48:1020–1026

    Article  PubMed  CAS  Google Scholar 

  • Vann DC (1995) Electroporation-based transformation of freshly harvested conidia of Neurospora crassa. Fungal Genet Newsl A 42:53

    Google Scholar 

  • Westergaard M, Mitchell HK (1947) Neurospora V. A synthetic medium favoring sexual reproduction. Am J Bot 34:573

    Article  Google Scholar 

  • Wickham H (2009) Introduction. In: ggplot2. Springer, pp 1–7

  • Wu C-Y, Bird AJ, Chung LM, Newton MA, Winge DR, Eide DJ (2008) Differential control of Zap1-regulated genes in response to zinc deficiency in Saccharomyces cerevisiae. BMC Genom 9:1

    Article  CAS  Google Scholar 

  • Wu YH, Frey AG, Eide DJ (2011) Transcriptional regulation of the Zrg17 zinc transporter of the yeast secretory pathway. Biochem J 435:259–266

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Yuan DS (2000) Zinc-regulated genes in Saccharomyces cerevisiae revealed by transposon tagging. Genetics 156:45–58

    PubMed  PubMed Central  CAS  Google Scholar 

  • Zhao H, Eide D (1996) The ZRT2 gene encodes the low affinity zinc transporter in Saccharomyces cerevisiae. J Biol Chem 271:23203–23210

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

Funding for this research project was provided by Science and Engineering Research Board (SERB), New Delhi (Grant no YSS/2014/000174). A.T. is supported by SERB Start-up Grant (Young Scientist).

Author information

Authors and Affiliations

  1. Department of Bioscience and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, Assam, 781039, India

    Anand Tiwari, Serena Daniel Ngiilmei & Ranjan Tamuli

Authors
  1. Anand Tiwari
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Serena Daniel Ngiilmei
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ranjan Tamuli
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Anand Tiwari.

Additional information

Communicated by M. Kupiec.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 1080 KB)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Tiwari, A., Ngiilmei, S.D. & Tamuli, R. The NcZrg-17 gene of Neurospora crassa encodes a cation diffusion facilitator transporter required for vegetative development, tolerance to endoplasmic reticulum stress and cellulose degradation under low zinc conditions. Curr Genet 64, 811–819 (2018). https://doi.org/10.1007/s00294-017-0794-4

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00294-017-0794-4

Keywords

Search

Navigation