Mitosis, gamma-tubulin function, cell cycle regulation and fungal secondary metabolites.
We currently have two major research projects. The first is on γ-tubulin and its role in cell cycle regulation. This project is funded by an R01 grant from NIGMS. We are using the filamentous fungus Aspergillus nidulans as our experimental organism for this project.
My lab discovered γ-tubulin and helped to uncover its role in microtubule nucleation. More recent work with γ-tubulin mutants has revealed that γ-tubulin has an important but poorly understood role in mitotic regulation independent of its role in microtubule nucleation. We are now attempting to understand this role (or these roles).
One of our approaches has been to tag mitotic regulatory proteins with fluorescent proteins and observe them in living cells of γ-tubulin mutant strains. The rationale is that by understanding how γ-tubulin mutants alter the behavior of mitotic regulatory proteins, we can begin to understand the role(s) of γ-tubulin in mitotic regulation. In order to carry out this project in a timely fashion, we have developed a very efficient gene targeting system for A. nidulans. This has allowed us to tag nearly all of the known mitotic regulatory proteins in this organism. (As will be mentioned below, the technique itself is an important advance for the A. nidulans community and has consequences well beyond this project.) So far this approach has uncovered a surprising role for γ-tubulin in regulating the inactivation of the anaphase promoting complex/cyclosome in the G1 phase of the cell cycle. Recent work is turning up additional surprises and we are excited that this project will tell us a great deal about cell cycle regulation that will go well beyond the functions of γ-tubulin.
Our second project is to identify and characterize secondary metabolites from A. nidulans and the genes involved in their biosynthesis. Secondary metabolites are not essential for viability but are often used to compete with other organisms. The classic example, of course, is penicillin. The biological activities of secondary metabolites are often medically useful and about half of the known fungal secondary metabolites have activities that are potentially useful. There are, of course, side effects and other problems so that most never make it into the pharmacopoeia, but there are some very notable successes such as lovastatin, a compound produced by Aspergillus terreus.
The sequencing of the genomes of species of Aspergillus revealed that they have the potential (based on the number of polyketide synthase and non-ribosomal peptide synthetase genes in the genome) to produce many more secondary metabolites than were known. In addition, the genes that encode fungal secondary metabolite biosynthesis genes are generally clustered, such that, for example, all of the genes involved in penicillin biosynthesis are adjacent in the genome. The sequence of the A. nidulans genome revealed that this organism had almost 40 more secondary metabolism gene clusters than known secondary metabolites. The gene targeting procedure we have developed for A. nidulans allows us to delete genes or replace their promoters with ease. This should allow us, in principle, to obtain expression of cryptic secondary metabolism pathways and, through deletion analysis, define the genes responsible for the synthesis of each secondary metabolite.
We have recently been awarded a program project grant from the NIGMS to support this work. I am the overall PI and Dr. Clay Wang at the University of Southern California and Dr. Nancy Keller at the University of Wisconsin are the other PI's. Dr. Wang is a natural products chemist and Dr. Keller studies expression of secondary metabolism genes. This work is going very well. We have discovered five compounds new to science and at least a half dozen new to A. nidulans in the few months since the grant began. In cases where the compounds were known to exist, we have been able to determine their biosynthetic pathways for the first time. We have also made progress in understanding chromatin-level regulation of secondary metabolism genes. This approach promises to revolutionize natural products chemistry.
- Liu, Ting, Sanchez, James, Chiang, Yi-Ming, Oakley, Berl R. and Wang, Clay C. C. (2014) Rational domain swaps reveal insights about chain length control by ketosynthase domains in fungal non-reducing polyketide synthases. Org. Lett. 16:1676-1679.
- Paranjape, Smita R., Chiang, Yi-Ming, Sanchez, James F., Entwistle, Ruth, Wang, Clay C. C., Oakley, Berl R. and Gamblin, T. Chris. (2014) Inhibition of tau aggregation by three Aspergillus nidulans secondary metabolites: 2,ω-dihydroxyemodin, asperthecin and asperbenzaldehyde. Planta Med. 80:77-85.
- Chiang, Yi-Ming, Wang, Clay C. C. and Oakley Berl R. (2014) Analyzing fungal secondary metabolite genes and gene clusters. In Natural Products: Discourse, Diversity & Design. (Edited by A. Osbourn, R. Goss and G. Carter), John Wiley & Sons, Hoboken, N.J. pp 175-193.
- Yaegashi, Junko, Oakley, B. R. and Wang, Clay C. C. (2014) Recent advances in genome mining of secondary metabolite biosynthetic gene clusters and the development of heterologous expression systems in Aspergillus nidulans. J. Ind. Microbiol. Biotechnol. 41:433-442.
- Oakley, Berl R. (2013) Aspergillus nidulans. In Encyclopedia of Genetics, 2nd edition (Edited by S. Maloy and K. Hughes), Academic Press, San Diego, CA. pp 212-215.
- Basha, Anwer, Basha, Fatima, Ali, Syed Kashif, Hanson, Paul R., Mitscher, Lester A. and Oakley, Berl R. (2013) Recent progress in the chemotherapy of human fungal diseases, Emphasis on 1,3-β-glucan synthase and chitin synthase inhibitors. Curr. Med. Chem. 20, 4859-4887.
- Yaegashi, Junko, Praseuth M. B., Tyan, S.-W. Sanchez, J. F., Entwistle, R., Chiang, Y.-M., Oakley, B. R. and Wang, C. C. C. (2013) Molecular genetic characterization of the biosynthesis cluster of a prenylated isoindolinone alkaloid aspernidine A in A. nidulans. Org. Lett. 15:2862-2865.
- Chiang, Yi-Ming, Oakley, C. Elizabeth, Ahuja, Manmeet, Entwistle, Ruth, Schultz, Aric, Chang, Shu-Lin, Sung, Calvin T., Wang, Clay C. C. and Oakley, Berl R. (2013) An efficient system for heterologous expression of secondary metabolite genes in Aspergillus nidulans. J. Am. Chem. Soc. 135:7720-7731.
- Yeh, H. H., Chang, S. L., Chiang, Y. M., Bruno, K. S., Oakley, B. R., Wu, T. K. and Wang, C. C. C. (2013). Engineering fungal nonreducing polyketide synthase by heterologous expression and domain swapping. Org Lett. 15:756-759.
- Guo, C.-J., Knox, B., Chiang, Y.-M., Lo, H.-C., Sanchez, J., Lee, K.-H., Oakley, B., Bruno, K., and Wang, C. (2012). Molecular genetic characterization of a cluster in A. terreus for biosynthesis of the meroterpenoid terretonin. Org. Lett. 14:5684-5687.
- Edgerton-Morgan, Heather and Oakley, Berl R. (2012) γ-Tubulin plays a key role in inactivating APC/CCdh1 at the G1/S boundary. J. Cell Biol. 198:785-791.
- Oakley, C. Elizabeth, Edgerton-Morgan, Heather, and Oakley, Berl R. (2012) Tools for Manipulation of Secondary Metabolism Pathways: Rapid Promoter Replacements and Gene Deletions in Aspergillus nidulans. In: Methods in Molecular Biology (Edited by N. Keller and G. Turner), Humana Press, Clifton, N. J. 944:143-161.
- Ahuja, Manmeet, Chiang, Yi-Ming, Chang, Shu-Lin, Praseuth, Mike B., Entwistle, Ruth, Sanchez, James F., Lo, Hsien-Chun, Oakley, Berl R. and Wang, Clay. C. C. (2012). Illuminating the diversity of aromatic polyketide synthases in Aspergillus nidulans. J. Am. Chem. Soc. 134, 8212-8221.
- Soukup, Alexandra A., Chaing, Yi-Ming, Bok, Jin Woo, Reyes-Dominguez, Yazmid, Oakley, Berl R., Wang, Clay C. C., Strauss, Joseph, and Keller Nancy P. (2012) Overexpression of the Aspergillus nidulans histone 4 acetyltransferase EseA increases activation of secondary metabolite production. Mol. Microbiol. 86:314-330.
- Sanchez, James F., Entwistle, Ruth, Corcoran, David, Oakley, Berl R., and Wang, Clay C. C. (2012) Identification and molecular genetic analysis of the cichorine cluster in Aspergillus nidulans. Med. Chem. Commun. 3:997-1002.
- Yeh, Hsu-Hua, Chiang, Yi-Ming, Entwistle, Ruth, Ahuja, Manmeet, Lee, Kuan-Han, Bruno, Kenneth, Wu, Tung-Kung, Oakley Berl. R. and Wang, Clay C. C. (2012) Molecular genetic analysis reveals that a nonribosomal peptide synthase-like (NRPS-like) gene in Aspergillus nidulans is responsible for microperfuranone biosynthesis. Appl. Microbiol. Biotechnol. 96:739-748.
- Lo, Hsien-Chun, Entwistle, Ruth, Guo, Chun-Jun, Ahuja, Manmeet, Ahuja, Manmeet, Szewczyk, Edyta, Hung, Jui-Hsiang, Chiang, Yi-Ming, Oakley, Berl R. and Wang, Clay C. C. (2012) Two separate gene clusters encode the biosynthetic pathway for the meroterpenoids, austinol and dehydroaustinol in Aspergillus nidulans. J. Am. Chem. Soc. 134, 4709-4720.
- Somoza, Amber D., Lee, Kuan-Han, Chiang, Yi-Ming, Oakley, Berl R. and Wang, Clay C. C. (2012) Reengineering an azaphilone biosynthesis pathway in Aspergillus nidulans to create lipoxygenase inhibitiors. Org. Lett. 14:972-975.
- Taheri-Talesh, Naimeh, Xiong, Yi and Oakley, Berl R. (2012) The functions of myosin II and myosin V homologs in tip growth and septation in Aspergillus nidulans. PLoS One 7, e31218.
- Liu, Ting, Chiang, Yi-Ming, Somoza, Amber D., Oakley, Berl R. and Wang, Clay C. C. (2011) Engineering of an “unnatural” natural product by swapping polyketide synthase domains in Aspergillus nidulans. J. Am. Chem. Soc. 133, 13314-13316.
- Szewczyk, Edyta and Oakley Berl R. (2011) Microtubule dynamics in mitosis in Aspergillus nidulans. Fungal Genet. Biol. 48, 998-999.
- Sanchez, James. F., Entwistle, Ruth, Hung, Jui-Hsiang, Yaegashi, Junko, Jain, Sofina, Chiang, Yi-Ming, Wang, Clay C. C. and Oakley, Berl R. (2011) Genome-based deletion analysis uncovers the prenyl xanthone biosynthesis pathway in Aspergillus nidulans. J. Am. Chem. Soc. 133, 4010-4017.
- Giles, Steve S., Soukup, Alexandra A., Lauer, Carrie, Shaaban, Mona, Lin, Alexander, Oakley, Berl R., Wang, Clay C. C. and Keller, Nancy P. (2011) Cryptic Aspergillus nidulans antimicrobials. Appl. Environ. Microbiol. 77, 3669-3675.
- Palmer, Jonathan M., Mallaredy, Sandeep, Perry, Dustin W., Sanchez, James F., Thiesen, Jeffrey M., Szewczyk, Edyta, Oakley, Berl R., Wang, Clay C. C., Keller, Nancy P., and Mirabito, Peter M. (2010) Telomere position effect is regulated by heterochromatin associated proteins and NkuA in Aspergillus nidulans. Microbiology 156, 3522-3531.
- Chiang, Yi-Ming, Chang, Shu-Lin, Oakley, Berl R. and Wang, Clay C. C. (2010). Recent advances in awakening silent biosynthetic gene clusters in microorganisms. Curr. Opin. Chem. Biol. 15,1-7.
- Nayak, Tania, Edgerton-Morgan, Heather, Horio, Tetsuya, Xiong, Yi, De Souza, Colin P., Osmani, Stephen A. and Oakley, Berl R. (2010) γ-Tubulin regulates the anaphase-promoting complex/cyclosome during interphase. J. Cell Biol. 190, 317-330.
- Chiang, Yi-Ming, Oakley, Berl R., Keller, Nancy P. and Wang, Clay C. C. (2010). Unraveling polyketide synthesis in members of the genus Aspergillus. Appl. Microbiol. Biotechnol. 86. 1719-1736.
- Chiang, Yi-Ming, Szewczyk, Edyta, Davidson, Ashley D., Entwistle, Ruth, Keller, Nancy P., Wang, Clay C. C. and Oakley, Berl R. (2010) Genetic characterization of the monodictyphenone gene cluster in Aspergillus nidulans. Appl. Environ. Microbiol. 76, 2067-2074.
- Sanchez, James F., Chiang, Yi-Ming, Szewczyk, Edyta, Davidson, Ashley D., Ahuja, Manmeet, Oakley, C. Elizabeth, Bok, Jin Woo, Keller, Nancy, Oakley, Berl R. and Wang, Clay C. C. (2010) Molecular genetic analysis of the orsellinic acid/F-9775 biosynthetic gene cluster in Aspergillus nidulans. Mol. Biosyst. 6, 587-593.
- Xiang, X. and Oakley, Berl R. (2010). The Cytoskeleton in Filamentous Fungi. In: Cellular and Molecular Biology of Filamentous Fungi (Edited by K Borkovich and D. Ebbole), ASM Press. pp 209-223.
Selected Earlier Publications
- Szewczyk, E., Nayak, T., Oakley, C. E., Edgerton, H., Xiong, Y., Taheri-Talesh, N., Osmani, S. A., and Oakley, B. R. (2006). Fusion PCR and gene targeting in Aspergillus nidulans. Nat. Protoc. 1, 3111–3120.
- Nayak, Tania, Szewczyk, Edyta, Oakley, C. Elizabeth, Osmani, Aysha, Ukil, Leena, Murray, Sandra L., Hynes, M. J., Osmani, Stephen A., and Oakley, B. R. (2006). A versatile and efficient gene-targeting system for Aspergillus nidulans. Genetics 172, 1557–1566.
- Galagan, J. E. et al. (2005). Sequencing of Aspergillus nidulans and comparative analysis with A. fumigatus and A. oryzae. Nature 438, 1105–1115.
- Horio, Tetsuya, and Oakley, B. R. (2005). The role of microtubules in rapid hyphal tip growth of Aspergillus nidulans. Mol. Biol. Cell 16, 918–926.
- Prigozhina, N. L., Oakley, C. E., Lewis, A., Nayak, Tania, Osmani, Stephen A., and Oakley, B. R. (2004). γ-Tubulin plays an essential role in the coordination of mitotic events. Mol. Biol. Cell 15, 1374–1386.
- Jung, M. Katherine, Prigozhina, Natalie, Oakley, C. E., Nogales, E., and Oakley, B. R. (2001). Alanine-scanning mutagenesis of Aspergillus γ-tubulin yields diverse and novel phenotypes. Mol. Biol. Cell 12, 2119–2136.
- Khodjakov, A., Cole, R. W., Oakley, B. R., and Rieder, C. L. (2000). Centrosome-independent mitotic spindle formation in vertebrates. Curr Biol 10, 59–67.
- Horio, T., and Oakley, B. R. (1992). Human γ tubulin functions in fission yeast.
- Zheng, Yixian, Jung, M. Katherine, and Oakley, B. R. (1991). γ-tubulin is present in Drosophila melanogaster and Homo sapiens and is associated with the centrosome. Cell 65, 817–823.
- Oakley, Berl R., Oakley, C. Elizabeth, Yoon, Yisang, M, Jung, K. (1990). γ tubulin is a component of the spindle-pole-body that is essential for microtubule function in Aspergillus nidulans. Cell 61, 1289–1301.
- Oakley, C. Elizabeth, and Oakley, B. R. (1989). Identification of γ-tubulin, a new member of the tubulin superfamily encoded by mipA gene of Aspergillus nidulans. Nature 338, 662–664.
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