- Email: [email protected]
Fungi learn how to surf big waves
JID:PLREV AID:472 /DIS
[m3SC+; v 1.191; Prn:31/03/2014; 13:56] P.1 (1-2)
Available online at www.sciencedirect.com
ScienceDirect Physics of Life Reviews ••• (••••) •••–••• www.elsevier.com/locate/plrev
Comment
Fungi learn how to surf big waves Comment on “Physical methods for genetic transformation of fungi and yeast” by Ana Leonor Rivera, Denis Magaña-Ortíz, Miguel Gómez-Lim, Francisco Fernández, Achim M. Loske Fausto Arellano-Carbajal Facultad de Ciencias Naturales, Universidad Autónoma de Querétaro, Av. de las Ciencias S/N, Juriquilla, Querétaro, Qro., México C.P. 76230, Mexico Received 7 March 2014; accepted 11 March 2014
Communicated by M. Frank-Kamenetskii
Fungi are among the most widely distributed organisms on Earth, with around 1.5 to 5 million of species of fungi estimated to date, and many of these are of great environmental, industrial, agricultural and medical importance [1]. In the genome race, the honor to be the first fully-sequenced eukaryotic genome fell on a fungus, Saccharomyces cerevisae [2], while metazoans had yet to wait for two years to be sequenced [3]. After Saccharomyces, 21 fungi have been sequenced and another thousand are now on the way [4]. Taking into account their relevance to different areas of applied science, having an efficient and reliable genetic transformation protocol is of essence [5]. Attempts for genetic transformation have been performed for nearly a century, since early work by Griffith who described genetic transformation of Pneumococcus [6], a feat successfully mirrored in S. cerevisae in 1960 by Oppenourt [7]. However, other research groups were unable to repeat these procedures. Finally, transformation of a filamentous fungus, Neurospora crassa, was achieved in 1979 [8], and in 1981 transformation of yeast was accomplished [9]. At the start of the 21st century, fungal transformation methods have been developed for all major fungal groups, albeit with varying efficiencies [10]. Currently there are two types of methods typically used for genetic transformation of fungi, namely biological and physical methods. The former are based on treatment of protoplasts with polyethylene glycol and enzymes, or on Agrobacterium tumefacinens. The latter are mainly based on electroporation and biolistics, or, alternatively, agitation with glass beads and vacuum infiltration. However, be they biological or physical, most available fungal transformation protocols are inefficient, laborious and have low reproducibility. These caveats, and the fact that transformation has not yet been accomplished for most of the more than one million fungal species that exist, highlight the importance of developing novel methodologies and of thinking, if you will, “outside of the box”. In the review article by Rivera et al. [11], a process termed “shock wave-mediated transformation” is presented as a novel alternative which the authors suggest will be safer, simpler and more efficient [12]. This novel method to transform fungi uses underwater shock waves and has already been used successfully to transform Escherichia coli, Pseudomonas aeruginosa and Salmonella typhimurium [13–15]. Shock waves are produced by a piezoelectric DOI of original article: http://dx.doi.org/10.1016/j.plrev.2014.01.007. http://dx.doi.org/10.1016/j.plrev.2014.03.003 1571-0645/© 2014 Elsevier B.V. All rights reserved.
JID:PLREV AID:472 /DIS
2
[m3SC+; v 1.191; Prn:31/03/2014; 13:56] P.2 (1-2)
F. Arellano-Carbajal / Physics of Life Reviews ••• (••••) •••–•••
shock wave generator, designed to expose fungal conidia to heterologous DNA. According to the authors previously published experiments, to date, three fungi of industrial importance (Aspergillus niger, Trichoderma reesei and Phanerochaete chrysosporium), and one phytopathogenic fungus (Fusarium oxysporum) have been successfully transformed using this device [12]. The advantages of using shock wave mediated transformation are multifaceted. On one hand, there is a budget benefit, as enzymes are not required, and as transformation efficiency is reportedly between two and four orders of magnitude higher than current protocols [11], research results are likely to be impacted positively. Furthermore, shock wave transformation appears to be a quick, easy to perform and, above all, a reproducible method for fungal transformation. Perhaps more importantly, the procedure appears to be feasible to implement on the close to a million fungal species for which transformation has not yet been accomplished. Detractors might argue that implementation of this method must require an initial high investment as relatively expensive equipment is needed, an extremely valid argument in times marked by important cuts in funding for science. However, even this point can fare positively in an argument. To sequence a genome at the turn of the century was five to six orders of magnitude more expensive than what it costs to do so now. This massive reduction is due mainly to the cost of the necessary equipment and reagents. If shock wave transformation becomes a more widely used procedure, I would venture to predict that the cost of the now expensive equipment will be reduced in time, making it feasible to purchase for a laboratory, department or institute. In their review article, Rivera et al. [11] have described and contrasted the different physical methods to transform fungi and have underscored the methodological advantages of their novel shock wave mediated transformation. The method seems extremely promising, and I can only hope that these new waves reach metazoan shores very soon. Perhaps we will finally see the humble worm learning how to surf big waves together with fungi. Acknowledgements Fausto Arellano-Carbajal is supported by funds from PROMEP-SEP (Mexico) (grant number PROMEP-SEP 40710884) and FOFI-UAQ (Mexico) (grant number FOFI-UAQ 20101169). References [1] Blackwell M. The fungi: 1, 2, 3 . . . 5.1 million species?. Am J Bot 2011;98:426–38. [2] Goffeau A, Barrell BG, Bussey H, Davis RW, Dujon B, Feldmann H, et al. Life with 6000 genes. Science 1996;274:563–7. [3] C. elegans Sequencing Consortium. Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 1998;282:2012–8. [4] http://1000.fungalgenomes.org/home/. [5] Utermark J, Karlovsky P. Genetic transformation of filamentous fungi by Agrobacterium tumefaciens. Nat Protoc 2008 [online]. http://dx.doi.org/10.1038/nprot.2008.83. [6] Griffith F. The significance of pneumococcal types. J Hyg 1928;27:113–59. [7] Oppenoorth WFF. Modification of the heredity character of yeast by ingestion of cell free extracts. Antonie Van Leeuwenhoek 1960;26:129–68. [8] Case ME, Schweizer M, Kushner SR, Giles NH. Efficient transformation of Neurospora crassa by utilizing hybrid plasmid DNA. Proc Natl Acad Sci USA 1979;76:5259–63. [9] Kimura A, Arima A, Murata K. Biofunctional change in yeast cell surface on treatment with Triton X-100. Agric Biol Chem 1981;45:2627–9. [10] Shigeyuki Kawai S, Wataru Hashimoto W, Kousaku Murata K. Transformation of Saccharomyces cerevisiae and other fungi. Methods and possible underlying mechanism. Bioeng Bugs 2010;6:395–403. [11] Rivera AL, Magaña-Ortíz D, Gómez-Lim M, Fernández F, Loske AM. Physical methods for genetic transformation of fungi and yeast. Phys Life Rev 2014. http://dx.doi.org/10.1016/j.plrev.2014.01.007 [in this issue]. [12] Magaña-Ortíz D, Coconi-Linares N, Ortiz-Vazquez E, Fernández F, Loske AM, Gómez-Lim MA. A novel and highly efficient method for genetic transformation of fungi employing shock waves. Fungal Genet Biol 2013;56:9–16. [13] Jagadeesh G, Nataraja KN. Udayakumar M. Shock waves can enhance bacterial transformation with plasmid DNA. Curr Sci India 2004;87:734–5. [14] Divya Prakash G, Anish RV, Jagadeesh G, Chakravortty D. Bacterial transformation using micro-shock waves. Anal Biochem 2011;419:292–301. [15] Loske AM, Campos-Guillen J, Fernández F, Castaño-Tostado E. Enhanced shock wave-assisted transformation of Escherichia coli. Ultrasound Med Biol 2011;37:502–10.
[m3SC+; v 1.191; Prn:31/03/2014; 13:56] P.1 (1-2)
Available online at www.sciencedirect.com
ScienceDirect Physics of Life Reviews ••• (••••) •••–••• www.elsevier.com/locate/plrev
Comment
Fungi learn how to surf big waves Comment on “Physical methods for genetic transformation of fungi and yeast” by Ana Leonor Rivera, Denis Magaña-Ortíz, Miguel Gómez-Lim, Francisco Fernández, Achim M. Loske Fausto Arellano-Carbajal Facultad de Ciencias Naturales, Universidad Autónoma de Querétaro, Av. de las Ciencias S/N, Juriquilla, Querétaro, Qro., México C.P. 76230, Mexico Received 7 March 2014; accepted 11 March 2014
Communicated by M. Frank-Kamenetskii
Fungi are among the most widely distributed organisms on Earth, with around 1.5 to 5 million of species of fungi estimated to date, and many of these are of great environmental, industrial, agricultural and medical importance [1]. In the genome race, the honor to be the first fully-sequenced eukaryotic genome fell on a fungus, Saccharomyces cerevisae [2], while metazoans had yet to wait for two years to be sequenced [3]. After Saccharomyces, 21 fungi have been sequenced and another thousand are now on the way [4]. Taking into account their relevance to different areas of applied science, having an efficient and reliable genetic transformation protocol is of essence [5]. Attempts for genetic transformation have been performed for nearly a century, since early work by Griffith who described genetic transformation of Pneumococcus [6], a feat successfully mirrored in S. cerevisae in 1960 by Oppenourt [7]. However, other research groups were unable to repeat these procedures. Finally, transformation of a filamentous fungus, Neurospora crassa, was achieved in 1979 [8], and in 1981 transformation of yeast was accomplished [9]. At the start of the 21st century, fungal transformation methods have been developed for all major fungal groups, albeit with varying efficiencies [10]. Currently there are two types of methods typically used for genetic transformation of fungi, namely biological and physical methods. The former are based on treatment of protoplasts with polyethylene glycol and enzymes, or on Agrobacterium tumefacinens. The latter are mainly based on electroporation and biolistics, or, alternatively, agitation with glass beads and vacuum infiltration. However, be they biological or physical, most available fungal transformation protocols are inefficient, laborious and have low reproducibility. These caveats, and the fact that transformation has not yet been accomplished for most of the more than one million fungal species that exist, highlight the importance of developing novel methodologies and of thinking, if you will, “outside of the box”. In the review article by Rivera et al. [11], a process termed “shock wave-mediated transformation” is presented as a novel alternative which the authors suggest will be safer, simpler and more efficient [12]. This novel method to transform fungi uses underwater shock waves and has already been used successfully to transform Escherichia coli, Pseudomonas aeruginosa and Salmonella typhimurium [13–15]. Shock waves are produced by a piezoelectric DOI of original article: http://dx.doi.org/10.1016/j.plrev.2014.01.007. http://dx.doi.org/10.1016/j.plrev.2014.03.003 1571-0645/© 2014 Elsevier B.V. All rights reserved.
JID:PLREV AID:472 /DIS
2
[m3SC+; v 1.191; Prn:31/03/2014; 13:56] P.2 (1-2)
F. Arellano-Carbajal / Physics of Life Reviews ••• (••••) •••–•••
shock wave generator, designed to expose fungal conidia to heterologous DNA. According to the authors previously published experiments, to date, three fungi of industrial importance (Aspergillus niger, Trichoderma reesei and Phanerochaete chrysosporium), and one phytopathogenic fungus (Fusarium oxysporum) have been successfully transformed using this device [12]. The advantages of using shock wave mediated transformation are multifaceted. On one hand, there is a budget benefit, as enzymes are not required, and as transformation efficiency is reportedly between two and four orders of magnitude higher than current protocols [11], research results are likely to be impacted positively. Furthermore, shock wave transformation appears to be a quick, easy to perform and, above all, a reproducible method for fungal transformation. Perhaps more importantly, the procedure appears to be feasible to implement on the close to a million fungal species for which transformation has not yet been accomplished. Detractors might argue that implementation of this method must require an initial high investment as relatively expensive equipment is needed, an extremely valid argument in times marked by important cuts in funding for science. However, even this point can fare positively in an argument. To sequence a genome at the turn of the century was five to six orders of magnitude more expensive than what it costs to do so now. This massive reduction is due mainly to the cost of the necessary equipment and reagents. If shock wave transformation becomes a more widely used procedure, I would venture to predict that the cost of the now expensive equipment will be reduced in time, making it feasible to purchase for a laboratory, department or institute. In their review article, Rivera et al. [11] have described and contrasted the different physical methods to transform fungi and have underscored the methodological advantages of their novel shock wave mediated transformation. The method seems extremely promising, and I can only hope that these new waves reach metazoan shores very soon. Perhaps we will finally see the humble worm learning how to surf big waves together with fungi. Acknowledgements Fausto Arellano-Carbajal is supported by funds from PROMEP-SEP (Mexico) (grant number PROMEP-SEP 40710884) and FOFI-UAQ (Mexico) (grant number FOFI-UAQ 20101169). References [1] Blackwell M. The fungi: 1, 2, 3 . . . 5.1 million species?. Am J Bot 2011;98:426–38. [2] Goffeau A, Barrell BG, Bussey H, Davis RW, Dujon B, Feldmann H, et al. Life with 6000 genes. Science 1996;274:563–7. [3] C. elegans Sequencing Consortium. Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 1998;282:2012–8. [4] http://1000.fungalgenomes.org/home/. [5] Utermark J, Karlovsky P. Genetic transformation of filamentous fungi by Agrobacterium tumefaciens. Nat Protoc 2008 [online]. http://dx.doi.org/10.1038/nprot.2008.83. [6] Griffith F. The significance of pneumococcal types. J Hyg 1928;27:113–59. [7] Oppenoorth WFF. Modification of the heredity character of yeast by ingestion of cell free extracts. Antonie Van Leeuwenhoek 1960;26:129–68. [8] Case ME, Schweizer M, Kushner SR, Giles NH. Efficient transformation of Neurospora crassa by utilizing hybrid plasmid DNA. Proc Natl Acad Sci USA 1979;76:5259–63. [9] Kimura A, Arima A, Murata K. Biofunctional change in yeast cell surface on treatment with Triton X-100. Agric Biol Chem 1981;45:2627–9. [10] Shigeyuki Kawai S, Wataru Hashimoto W, Kousaku Murata K. Transformation of Saccharomyces cerevisiae and other fungi. Methods and possible underlying mechanism. Bioeng Bugs 2010;6:395–403. [11] Rivera AL, Magaña-Ortíz D, Gómez-Lim M, Fernández F, Loske AM. Physical methods for genetic transformation of fungi and yeast. Phys Life Rev 2014. http://dx.doi.org/10.1016/j.plrev.2014.01.007 [in this issue]. [12] Magaña-Ortíz D, Coconi-Linares N, Ortiz-Vazquez E, Fernández F, Loske AM, Gómez-Lim MA. A novel and highly efficient method for genetic transformation of fungi employing shock waves. Fungal Genet Biol 2013;56:9–16. [13] Jagadeesh G, Nataraja KN. Udayakumar M. Shock waves can enhance bacterial transformation with plasmid DNA. Curr Sci India 2004;87:734–5. [14] Divya Prakash G, Anish RV, Jagadeesh G, Chakravortty D. Bacterial transformation using micro-shock waves. Anal Biochem 2011;419:292–301. [15] Loske AM, Campos-Guillen J, Fernández F, Castaño-Tostado E. Enhanced shock wave-assisted transformation of Escherichia coli. Ultrasound Med Biol 2011;37:502–10.