1887

Abstract

Melanin is a black pigment widely distributed across the kingdoms, from bacterial to human. The filamentous fungus is a typical ‘black fungus’, which produces melanin in its hyphal and especially its asexual spore cell walls. Its biosynthesis follows the dihydroxynaphthalene (DHN) pathway with 1,8-DHN as an intermediate. Two genes, encoding a polyketide synthase () and a 1,3,8-trihydroxynaphthalene (THN) reductase (), along with a putative transcription factor, CmrA, comprise a small gene cluster. Here we show that CmrA controls the expression of and , but that it also controls the expression of a scytalone dehydratase encoding gene () located elsewhere in the genome. The regulatory function of CmrA was shown in a reporter assay system. CmrA was expressed in the filamentous fungus where it was able to induce the expression of a reporter construct under the control of the putative promoter. This suggests direct binding of CmrA to the promoter of in the heterologous system. Likewise, silencing of in led to white colonies due to the lack of melanin. In addition, hyphal diameter and spore morphology were changed in the mutant and the number of spores reduced. Silencing of and inhibition of melanin biosynthesis by tricyclazole largely phenocopied the effects of silencing, suggesting a novel regulatory function of melanin in morphogenetic pathways.

Funding
This study was supported by the:
  • BMBF (Award 0316052C)
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.079046-0
2014-09-01
2024-12-12
Loading full text...

Full text loading...

/deliver/fulltext/micro/160/9/1845.html?itemId=/content/journal/micro/10.1099/mic.0.079046-0&mimeType=html&fmt=ahah

References

  1. Aguirre J., Ríos-Momberg M., Hewitt D., Hansberg W. ( 2005). Reactive oxygen species and development in microbial eukaryotes. Trends Microbiol 13:111–118 [View Article][PubMed]
    [Google Scholar]
  2. Bechinger C., Giebel K. F., Schnell M., Leiderer P., Deising H. B., Bastmeyer M. ( 1999). Optical measurements of invasive forces exerted by appressoria of a plant pathogenic fungus. Science 285:1896–1899 [View Article][PubMed]
    [Google Scholar]
  3. Carzaniga R., Fiocco D., Bowyer P., O’Connell R. J. ( 2002). Localization of melanin in conidia of Alternaria alternata using phage display antibodies. Mol Plant Microbe Interact 15:216–224 [View Article][PubMed]
    [Google Scholar]
  4. Cho Y., Davis J. W., Kim K.-H., Wang J., Sun Q.-H., Cramer R. A. Jr, Lawrence C. B. ( 2006). A high throughput targeted gene disruption method for Alternaria brassicicola functional genomics using linear minimal element (LME) constructs. Mol Plant Microbe Interact 19:7–15 [View Article][PubMed]
    [Google Scholar]
  5. Cho Y., Srivastava A., Ohm R. A., Lawrence C. B., Wang K.-H., Grigoriev I. V., Marahatta S. P. ( 2012). Transcription factor Amr1 induces melanin biosynthesis and suppresses virulence in Alternaria brassicicola. PLoS Pathog 8:e1002974 [View Article][PubMed]
    [Google Scholar]
  6. Eisenman H. C., Casadevall A. ( 2012). Synthesis and assembly of fungal melanin. Appl Microbiol Biotechnol 93:931–940 [View Article][PubMed]
    [Google Scholar]
  7. Eliahu N., Igbaria A., Rose M. S., Horwitz B. A., Lev S. ( 2007). Melanin biosynthesis in the maize pathogen Cochliobolus heterostrophus depends on two mitogen-activated protein kinases, Chk1 and Mps1, and the transcription factor Cmr1. Eukaryot Cell 6:421–429 [View Article][PubMed]
    [Google Scholar]
  8. Heinekamp T., Thywißen A., Macheleidt J., Keller S., Valiante V., Brakhage A. A. ( 2012). Aspergillus fumigatus melanins: interference with the host endocytosis pathway and impact on virulence. Front Microbiol 3:440[PubMed]
    [Google Scholar]
  9. Hill T. W., Kafer E. ( 2001). Improved protocols for Aspergillus minimal medium: trace element and minimal medium salt stock solutions. Fungal Genet Newsl 48:20–21
    [Google Scholar]
  10. Howard R. J., Ferrari M. A., Roach D. H., Money N. P. ( 1991). Penetration of hard substrates by a fungus employing enormous turgor pressures. Proc Natl Acad Sci U S A 88:11281–11284 [View Article][PubMed]
    [Google Scholar]
  11. Hynes M. J., Murray S. L., Khew G. S., Davis M. A. ( 2008). Genetic analysis of the role of peroxisomes in the utilization of acetate and fatty acids in Aspergillus nidulans.. Genetics 178:1355–1369 [View Article][PubMed]
    [Google Scholar]
  12. Jacobson E. S., Hove E., Emery H. S. ( 1995). Antioxidant function of melanin in black fungi. Infect Immun 63:4944–4945[PubMed]
    [Google Scholar]
  13. Kawamura C., Tsujimoto T., Tsuge T. ( 1999). Targeted disruption of a melanin biosynthesis gene affects conidial development and UV tolerance in the Japanese pear pathotype of Alternaria alternata. Mol Plant Microbe Interact 12:59–63 [View Article][PubMed]
    [Google Scholar]
  14. Kimura N., Tsuge T. ( 1993). Gene cluster involved in melanin biosynthesis of the filamentous fungus Alternaria alternata.. J Bacteriol 175:4427–4435[PubMed]
    [Google Scholar]
  15. Lee J.-K., Jung H.-M., Kim S.-Y. ( 2003). 1,8-dihydroxynaphthalene (DHN)-melanin biosynthesis inhibitors increase erythritol production in Torula corallina, and DHN-melanin inhibits erythrose reductase. Appl Environ Microbiol 69:3427–3434 [View Article][PubMed]
    [Google Scholar]
  16. Liu G. Y., Nizet V. ( 2009). Color me bad: microbial pigments as virulence factors. Trends Microbiol 17:406–413 [View Article][PubMed]
    [Google Scholar]
  17. Liu P. C., Thiele D. J. ( 2001). Novel stress-responsive genes EMG1 and NOP14 encode conserved, interacting proteins required for 40S ribosome biogenesis. Mol Biol Cell 12:3644–3657 [View Article][PubMed]
    [Google Scholar]
  18. Mander G. J., Wang H., Bodie E., Wagner J., Vienken K., Vinuesa C., Foster C., Leeder A. C., Allen G. & other authors ( 2006). Use of laccase as a novel, versatile reporter system in filamentous fungi. Appl Environ Microbiol 72:5020–5026 [View Article][PubMed]
    [Google Scholar]
  19. Martel C. M., Parker J. E., Warrilow A. G., Rolley N. J., Kelly S. L., Kelly D. E. ( 2010). Complementation of a Saccharomyces cerevisiae ERG11/CYP51 (sterol 14α-demethylase) doxycycline-regulated mutant and screening of the azole sensitivity of Aspergillus fumigatus isoenzymes CYP51A and CYP51B. Antimicrob Agents Chemother 54:4920–4923 [View Article][PubMed]
    [Google Scholar]
  20. Nakayashiki H., Hanada S., Nguyen B. Q., Kadotani N., Tosa Y., Mayama S. ( 2005). RNA silencing as a tool for exploring gene function in ascomycete fungi. Fungal Genet Biol 42:275–283 [View Article][PubMed]
    [Google Scholar]
  21. Pruß S., Fetzner R., Seither K., Herr A., Pfeiffer E., Metzler M., Lawrence C. B., Fischer R. ( 2014). Role of the Alternaria alternata blue-light receptor LreA (white-collar 1) in spore formation and secondary metabolism. Appl Environ Microbiol 80:2582–2591 [View Article][PubMed]
    [Google Scholar]
  22. Saha D., Fetzner R., Burkhardt B., Podlech J., Metzler M., Dang H., Lawrence C. B., Fischer R. ( 2012). Identification of a polyketide synthase required for alternariol (AOH) and alternariol-9-methyl ether (AME) formation in Alternaria alternata.. PLoS ONE 7:e40564 [View Article][PubMed]
    [Google Scholar]
  23. Stringer M. A., Dean R. A., Sewall T. C., Timberlake W. E. ( 1991). Rodletless, a new Aspergillus developmental mutant induced by directed gene inactivation. Genes Dev 5:1161–1171 [View Article][PubMed]
    [Google Scholar]
  24. Toews M. W., Warmbold J., Konzack S., Rischitor P., Veith D., Vienken K., Vinuesa C., Wei H., Fischer R. ( 2004). Establishment of mRFP1 as a fluorescent marker in Aspergillus nidulans and construction of expression vectors for high-throughput protein tagging using recombination in vitro (GATEWAY). Curr Genet 45:383–389 [View Article][PubMed]
    [Google Scholar]
  25. Tseng M. N., Chung P. C., Tzean S. S. ( 2011). Enhancing the stress tolerance and virulence of an entomopathogen by metabolic engineering of dihydroxynaphthalene melanin biosynthesis genes. Appl Environ Microbiol 77:4508–4519 [View Article][PubMed]
    [Google Scholar]
  26. Tsuge T., Harimoto Y., Akimitsu K., Ohtani K., Kodama M., Akagi Y., Egusa M., Yamamoto M., Otani H. ( 2013). Host-selective toxins produced by the plant pathogenic fungus Alternaria alternata.. FEMS Microbiol Rev 37:44–66 [View Article][PubMed]
    [Google Scholar]
  27. Tsuji G., Kenmochi Y., Takano Y., Sweigard J., Farrall L., Furusawa I., Horino O., Kubo Y. ( 2000). Novel fungal transcriptional activators, Cmr1p of Colletotrichum lagenarium and Pig1p of Magnaporthe grisea, contain Cys2His2 zinc finger and Zn(II)2Cys6 binuclear cluster DNA-binding motifs and regulate transcription of melanin biosynthesis genes in a developmentally specific manner. Mol Microbiol 38:940–954 [View Article][PubMed]
    [Google Scholar]
  28. Waring R. B., May G. S., Morris N. R. ( 1989). Characterization of an inducible expression system in Aspergillus nidulans using alcA and tubulin-coding genes. Gene 79:119–130 [View Article][PubMed]
    [Google Scholar]
  29. Yelton M. M., Hamer J. E., Timberlake W. E. ( 1984). Transformation of Aspergillus nidulans by using a trpC plasmid. Proc Natl Acad Sci U S A 81:1470–1474 [View Article][PubMed]
    [Google Scholar]
/content/journal/micro/10.1099/mic.0.079046-0
Loading
/content/journal/micro/10.1099/mic.0.079046-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF
This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error