Skip to main content
SpringerLink
Log in

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

  • Microbial Genetics, Genomics and Molecular Biology
  • Published:
Journal of Microbiology Aims and scope Submit manuscript

Abstract

Phospholipase C1 (PLC1), secretory phospholipase A2 (sPLA2) and Ca2+/H+ exchanger proteins regulate calcium signaling and homeostasis in eukaryotes. In this study, we investigate functions for phospholipase C1 (plc-1), sPLA2 (splA2) and a Ca2+/H+ exchanger (cpe-1) in the filamentous fungus Neurospora crassa. The Δplc-1, ΔsplA2, and Δcpe-1 mutants exhibited a growth defect on medium supplemented with the divalent ionophore A23187, suggesting that these genes might play a role in regulation of cytosolic free Ca2+ concentration ([Ca2+]c) in N. crassa. The strains lacking plc-1, splA2, and cpe-1 possessed higher carotenoid content than wild type at 8°C, 22°C, and 30°C, and showed increased ultraviolet (UV)-survival under conditions that induced carotenoid accumulation. Moreover, Δplc-1, ΔsplA2, and Δcpe-1 mutants showed reduced survival rate under hydrogen peroxide-induced oxidative stress and induced thermotolerance after exposure to heat shock temperatures. Thus, this study revealed multiple cellular roles for plc-1, splA2, and cpe-1 genes in regulation of [Ca2+]c, carotenoid accumulation, survival under stress conditions, and acquisition of thermotolerance induced by heat shock.

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

Similar content being viewed by others

References

  • Alcántara-Sánchez, F., Reynaga-Peña, C.G., Salcedo-Hernández, R., and Ruiz-Herrera, J. 2004. Possible role of ionic gradients in the apical growth of Neurospora crassa. Antonie van Leeuwenhoek 86, 301–311.

    Article  PubMed  Google Scholar 

  • Altschul, S.F., Gish, W., Miller, W., Myers, E.W., and Lipman, D.J. 1990. Basic local alignment search tool. J. Mol. Biol. 215, 403–410.

    Article  CAS  PubMed  Google Scholar 

  • Avalos, J., Luis, M., and Corrochano, L.M. 2013. Carotenoid Biosynthesis in Neurospora, pp. 227–241. In Kasbekar, D.P. McCluskey, K. (eds.), Neurospora: genomics and molecular biology. Caister Academic Press, Norfolk, UK.

  • Balestrieri, B., Maekawa, A., Xing, W., Gelb, M.H., Katz, H.R., and Arm, J.P. 2009. Group V secretory phospholipase A2 modulates phagosome maturation and regulates the innate immune response against Candida albicans. J. Immunol. 182, 4891–4898.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Berridge, M.J., Bootman, M.D., and Lipp, P. 1998. Calcium–a life and death signal. Nature 395, 645–648.

    Article  CAS  PubMed  Google Scholar 

  • Boilard, E., Lai, Y., Larabee, K., Balestrieri, B., Ghomashchi, F., Fujioka, D., Gobezie, R., Coblyn, J.S., Weinblatt, M.E., Massarotti, E.M., et al. 2010. A novel anti-inflammatory role for secretory phospholipase A2 in immune complex-mediated arthritis. EMBO Mol. Med. 2, 172–187.

    Article  PubMed Central  PubMed  Google Scholar 

  • Bootman, M.D., Collins, T.J., Peppiatt, C.M., Prothero, L.S., Mac-Kenzie, L., De Smet, P., Travers, M., Tovey, S.C., Seo, J.T., Berridge, M.J., et al. 2001. Calcium signalling–an overview. Semin. Cell Dev. Biol. 12, 3–10.

    Article  CAS  PubMed  Google Scholar 

  • Borkovich, K.A., Alex, L.A., Yarden, O., Freitag, M., Turner, G.E., Read, N.D., Seiler, S., Bell-Pedersen, D., Paietta, J., Plesofsky, N., et al. 2004. Lessons from the genome sequence of Neurospora crassa: tracing the path from genomic blueprint to multicellular organism. Microbiol. Mol. Biol. Rev. 68, 1–108.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bowman, B.J., Abreu, S., Margolles-Clark, E., Draskovic, M., and Bowman, E.J. 2011. Role of four calcium transport proteins, encoded by nca-1, nca-2, nca-3, and cax, in maintaining intracellular calcium levels in Neurospora crassa. Eukaryot. Cell 10, 654–661.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Cavazzini, D., Meschi, F., Corsini, R., Bolchi, A., Rossi, G.L., Einsle, O., and Ottonello, S. 2013. Autoproteolytic activation of a symbiosis-regulated truffle phospholipase A2. J. Biol. Chem. 288, 1533–1547.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Chae, S.W., Kim, J.M., Yun, Y.P., Lee, W.K., Kim, J.S., Kim, Y.H., Lee, K.S., Ko, Y.J., Lee, K.H., and Rha, H.K. 2007. Identification and analysis of the promoter region of the human PLC-δ4 gene. Mol. Biol. Rep. 34, 69–77.

    Article  CAS  PubMed  Google Scholar 

  • Chin, D. and Means, A R. 2000. Calmodulin: a prototypical calcium sensor. Trends Cell Biol. 10, 322–328.

    Article  CAS  PubMed  Google Scholar 

  • Choi, J., Kim, K.S., Rho, H.S., and Lee, Y.H. 2011. Differential roles of the phospholipase C genes in fungal development and pathogenicity of Magnaporthe oryzae. Fungal Genet. Biol. 48, 445–455.

    Article  CAS  PubMed  Google Scholar 

  • Chung, H.J., Kim, M.J., Lim, J.Y., Park, S.M., Cha, B.J., Kim, Y.H., Yang, M.S., and Kim, D.H. 2006. A gene encoding phosphatidyl inositol-specific phospholipase C from Cryphonectria parasitica modulates the lac1 expression. Fungal Genet. Biol. 43, 326–336.

    Article  CAS  PubMed  Google Scholar 

  • Clapham, D.E. 2007. Calcium signaling. Cell 131, 1047–1058.

    Article  CAS  PubMed  Google Scholar 

  • Cornelius, G. and Nakashima, H. 1987. Vacuoles play a decisive role in calcium homeostasis in Neurospora crassa. J. Gen. Microbiol. 133, 2341–2347.

    CAS  Google Scholar 

  • Davis, R.H. and De Serres, F.J. 1970. Genetic and microbial research techniques for Neurospora crassa. Methods Enzymol. 17, 79–143.

    Article  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  • Deka. R. and Tamuli, R. 2013. Neurospora crassa ncs-1, mid-1 and nca-2 double-mutant phenotypes suggest diverse interaction among three Ca2+-regulating gene products. J. Genet. 92, 559–563.

    Article  PubMed  Google Scholar 

  • Díaz-Sánchez, V., Estrada, A. F., Trautmann, D., Limón, M. C., Al-Babili, S., and Avalos, J. 2011. Analysis of al-2 mutations in Neurospora. PLoS ONE 6, e21948.

    Article  PubMed Central  PubMed  Google Scholar 

  • Estrada, A.F., Youssar, L., Scherzinger, D., Al-Babili, S., and Avalos, J. 2008. The ylo-1 gene encodes an aldehyde dehydrogenase responsible for the last reaction in the Neurospora carotenoid pathway. Mol. Microbiol. 69, 1207–1220.

    Article  CAS  PubMed  Google Scholar 

  • Felsenstein, J. 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39, 783–791.

    Article  Google Scholar 

  • Flick, J.S. and Thorner, J. 1993. Genetic and biochemical characterization of a phosphatidyl-inositol specific phospholipase C in Saccharomyces cerevisiae. Mol. Cell Biol. 13, 5861–5876.

    PubMed Central  CAS  PubMed  Google Scholar 

  • Galagan, J.E., Calvo, S.E., Borkovich, K.A., Selker, E.U., Read, N.D., Jaffe, D., FitzHugh, W., Ma, L.J., Smirnov, S., Purcell, S., et al. 2003. The genome sequence of the filamentous fungus Neurospora crassa. Nature 422, 859–868.

    Article  CAS  PubMed  Google Scholar 

  • Gavric, O., Becker dos Santos, D., and Griffiths, A. 2007. Mutation and divergence of the phospholipase C gene in Neurospora crassa. Fungal Genet. Biol. 44, 242–249.

    Article  CAS  PubMed  Google Scholar 

  • Guttery, D.S., Pittman, J.K., Freñal, K., Poulin, B., McFarlane, L.R., Slavic, K., Wheatley, S.P., Soldati-Favre, D., Krishna, S., Tewari, R., et al. 2013. The Plasmodium berghei Ca2+/H+ exchanger, PbCAX, is essential for tolerance to environmental Ca2+ during sexual development. PLoS Pathog. 9, e1003191.

  • Harding, R.W. 1974. The effect of temperature on photo-induced carotenoid biosynthesis in Neurospora crassa. Plant Physiol. 54, 142–147.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Harding, R.W., Huang, P.C., and Mitchell, H.K. 1969. Photochemical studies of the carotenoid biosynthetic pathway in Neurospora crassa. Arch. Biochem. Biophys. 129, 696–707.

    Article  CAS  PubMed  Google Scholar 

  • Kapoor, M., Sreenivasan, G.M., Goel, N., and Lewis, J. 1990. Development of thermotolerance in Neurospora crassa by heat shock and other stresses eliciting peroxidase induction. J. Bacteriol. 172, 2798–2801.

    PubMed Central  CAS  PubMed  Google Scholar 

  • Kapoor, M., Curle, C.A., and Runham, C. 1995. The hsp70 gene family of Neurospora crassa: cloning, sequence analysis, expression, and genetic mapping of the major stress-inducible member. J. Bacteriol. 177, 212–221.

    PubMed Central  CAS  PubMed  Google Scholar 

  • Kazmierczak, J., Kempe, S., and Kremer, B. 2013. Calcium in the early evolution of living systems: a biohistorical approach. Curr. Org. Chem. 17, 1738–1750.

    Article  CAS  Google Scholar 

  • Köhler, G.A., Brenot, A., Haas-Stapleton, E., Agabian, N., Deva, R., and Nigam, S. 2006. Phospholipase A2 and phospholipase B activities in fungi. Biochim. Biophys. Acta. 1761, 1391–1399.

    Article  PubMed Central  PubMed  Google Scholar 

  • Kumar, R. and Tamuli, R. 2014. Calcium/calmodulin-dependent kinases are involved in growth, thermotolerance, oxidative stress survival, and fertility in Neurospora crassa. Arch. Microbiol. 196, 295–305.

    Article  CAS  PubMed  Google Scholar 

  • Lev, S., Desmarini, D., Li, C., Chayakulkeeree, M., Traven, A., Sorrell, T.C., and Djordjevic, J.T. 2013. Phospholiase C of Crytococcus neoformans regulates homeostasis and virulence by providing inositol trisphosphate as a substrate for Arg1 Kinase. Infect. Immun. 81, 1245–1255.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Lew, R.R., Abbas, Z., Anderca, M.I., and Free, S.J. 2008. Phenotype of a mechanosensitive channel mutant, mid-1, in a filamentous fungus, Neurospora crassa. Eukaryot. Cell 7, 647–655.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Luque, E.M., Gutierrez, G., Navarro-Sampedro, L., Olmedo, M., Rodríguez-Romero, J., Ruger-Herreros, C., Tagua, V.G., and Corrochano, L.M. 2012. A relationship between carotenoid accumulation and the distribution of species of the fungus Neurospora in Spain. PLoS ONE 7, e33658.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • McCluskey, K., Wiest, A., and Plamann, M. 2010. The fungal genetics stock center: a repository for 50 years of fungal genetics research. J. Biosci. 35, 119–126.

    Article  CAS  PubMed  Google Scholar 

  • Murakami, M. and Kudo, I. 2002. Phospholipase A2. J. Biochem. 131, 285–292.

    Article  CAS  PubMed  Google Scholar 

  • Nakahama, T., Nakanishi, Y., Viscomi, A.R., Takaya, K., Kitamoto, K., Ottonello, S., and Arioka, M. 2010. Distinct enzymatic and cellular characteristics of two secretory phospholipases A2 in the filamentous fungus Aspergillus oryzae. Fungal Genet. Biol. 47, 318–331.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Nakashima, S., Wakatsuki, S., Yokoyama, T., Arioka, M., and Kitamoto, K. 2003. Identification and characterization of Scp15, a novel protein from Streptomyces coelicolor A3(2) with neuriteinducing activity in PC12 cells. Biosci. Biotechnol. Biochem. 67, 77–82

    Article  CAS  PubMed  Google Scholar 

  • Nelson, G., Kozlova-Zwinderman, O., Collis, A.J., Knight, M.R., Fincham, J.R., Stanger, C.P., Renwick, A., Hessing, J.G., Punt, P.J., Van den Hondel, C.A., et al. 2004. Calcium measurement in living filamentous fungi expressing codon-optimized aequorin. Mol. Microbiol. 52, 1437–1450.

    Article  CAS  PubMed  Google Scholar 

  • Nicholas, K.B. and Nicholas, H.B. 1997. GeneDoc: a tool for editing and annotating multiple sequence alignments. Distributed by the author. http://www.psc.edu/biomed/genedoc.

    Google Scholar 

  • Rho, H.S., Jeon, J., and Lee, Y.H. 2009. Phospholipase C-mediated calcium signalling is required for fungal development and pathogenicity in Magnaporthe oryzae. Mol. Plant Pathol. 10, 337–346.

    Article  CAS  PubMed  Google Scholar 

  • Rodriguez-Amaya, D.B. and Kimura, M. 2004. Harvest plus handbook for carotenoid analysis. Harvest Plus Technical Monograph 2. International Food Policy Research Institute (IFPRI) and International Center for Tropical Agriculture (CIAT), Washington, DC, USA.

    Google Scholar 

  • Rzhetsky, A. and Nei, M. 1992. Statistical properties of the ordinary least-squares, generalized least-squares, and minimum-evolution methods of phylogenetic inference. J. Mol. Evol. 35, 367–375.

    Article  CAS  PubMed  Google Scholar 

  • Schumacher, J., Viaud, M., Simon, A., and Tudzynski, B. 2008. The G alpha subunit BCG1, the phospholipase C (BcPLC1) and the calcineurin phosphatase co-ordinately regulate gene expression in the grey mould fungus Botrytis cinerea. Mol. Microbiol. 67, 1027–1050.

    Article  CAS  PubMed  Google Scholar 

  • Soragni, E., Bolchi, A., Balestrini, R., Gambaretto, C., Percudani, R., Bonfante, P., and Ottonello, S. 2001. A nutrient-regulated, dual localization phospholipase A(2) in the symbiotic fungus Tuber borchii. EMBO J. 20, 5079–5090.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Tamuli, R., Kumar, R., and Deka, R. 2011. Cellular roles of neuronal calcium sensor-1 and calcium/calmodulin-dependent kinases in fungi. J. Basic Microbiol. 51, 120–128.

    Article  CAS  PubMed  Google Scholar 

  • Tamuli, R., Kumar, R., Srivastava, D.A., and Deka, R. 2013. Calcium signaling, pp. 35–57. In Kasbekar, D.P. and McCluskey, K. (eds.), Neurospora: genomics and molecular biology. Caister Academic Press, Norfolk, UK.

  • Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M., and Kumar, S. 2011. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol. Biol. Evol. 28, 2731–2739.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Thompson, J.D., Gibson, T.J., Plewniak, F., Jeanmougin, F., and Higgins, D.G. 1997. The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res. 25, 4876–4882.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Tsai, H.C. and Chung, K.R. 2014. Calcineurin phosphatase and phospholipase C are required for developmental and pathological functions in the citrus fungal pathogen Alternaria alternata. Microbiology 160, 1453–1465.

    Article  CAS  PubMed  Google Scholar 

  • VogeL, H.J. 1964. Distribution of lysine pathways among fungi: evolutionary implications. Am. Naturalist 98, 435–446.

    Article  CAS  Google Scholar 

  • Wakatsuki, S., Arioka, M., Dohmae, N., Takio, K., Yamasaki, M., and Kitamoto, K. 1999. Characterization of a novel fungal protein, p15, which induces neuronal differentiation of PC12 cells. J Biochem. 126, 1151–1160. Erratum in: J Biochem. 127, 939.

    Article  CAS  PubMed  Google Scholar 

  • Wakatsuki, S., Yokoyama, T., Nakashima, S., Nishimura, A., Arioka, M., and Kitamoto, K. 2001. Molecular cloning, functional expression and characterization of p15, a novel fungal protein with potent neurite-inducing activity in PC12 cells. Biochim. Biophys. Acta 1522, 74–81.

    Article  CAS  PubMed  Google Scholar 

  • Yang, Q. and Borkovich, K.A. 1999. Mutational activation of a Gai causes uncontrolled proliferation of aerial hyphae and increased sensitivity to heat and oxidative stress in Neurospora crassa. Genetics 151, 107–117.

    PubMed Central  CAS  PubMed  Google Scholar 

  • Zalokar, M. 1954. Studies on biosynthesis of carotenoids in Neurospora crassa. Arch. Biochem. Biophys. 50, 71–80.

    Article  CAS  PubMed  Google Scholar 

  • Zelter, A., Bencina, M., Bowman, B.J., Yarden, O., and Read, N.D. 2004. A comparative genomic analysis of the calcium signaling machinery in Neurospora crassa, Magnaporthe grisea, and Saccharomyces cerevisiae. Fungal Genet. Biol. 41, 827–841.

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati, 781 039, India

    Ananya Barman & Ranjan Tamuli

Authors
  1. Ananya Barman
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ranjan Tamuli
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ranjan Tamuli.

Additional information

Supplemental material for this article may be found at http://www.springerlink.com/content/120956.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Barman, A., Tamuli, R. 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 (2015). https://doi.org/10.1007/s12275-015-4465-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12275-015-4465-1

Keywords

Search

Navigation