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Elucidating steroid alkaloid biosynthesis in Veratrum californicum: production of verazine in Sf9 cells |
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| Authors: | Megan M. Augustin Dan R. Ruzicka Ashutosh K. Shukla Jörg M. Augustin Courtney M. Starks Mark O'Neil‐Johnson Michael R. McKain Bradley S. Evans Matt D. Barrett Ann Smithson Gane Ka‐Shu Wong Michael K. Deyholos Patrick P. Edger J. Chris Pires James H. Leebens‐Mack David A. Mann Toni M. Kutchan |
| Affiliation: | 1. Donald Danforth Plant Science Center, St. Louis, MO, USA;2. Monsanto Company, St Louis, MO, USA;3. CSIR‐Central Institute of Medicinal and Aromatic Plants, Lucknow, Uttar Pradesh, India;4. Sequoia Sciences, St. Louis, MO, USA;5. Botanic Gardens and Parks Authority, Kings Park and Botanic Garden, West Perth 6005, WA, Australia;6. School of Plant Biology, University of Western Australia, Perth, WA, Australia;7. Department of Biological Sciences, University of Alberta, Edmonton, AB, Canada;8. Department of Medicine, University of Alberta, Edmonton, AB T6G 2E9, Canada;9. BGI‐Shenzhen, Build 11, Beishan Industrial Zone, Yantian District, Shenzhen, China;10. University of Alberta, Edmonton, AB T6G 2R3, Canada;11. Division of Biological Sciences, 311 Bond Life Sciences Center, University of Missouri, Columbia, MO, USA;12. Department of Plant and Microbial Biology, University of California, Berkeley, CA, USA;13. Franklin College of Arts and Sciences, University of Georgia, Athens, GA, USA;14. Infinity Pharmaceuticals, Cambridge, MA, USA;15. Cellular Dynamics International, Madison, WI, USA |
| Abstract: | Steroid alkaloids have been shown to elicit a wide range of pharmacological effects that include anticancer and antifungal activities. Understanding the biosynthesis of these molecules is essential to bioengineering for sustainable production. Herein, we investigate the biosynthetic pathway to cyclopamine, a steroid alkaloid that shows promising antineoplastic activities. Supply of cyclopamine is limited, as the current source is solely derived from wild collection of the plant Veratrum californicum. To elucidate the early stages of the pathway to cyclopamine, we interrogated a V. californicum RNA‐seq dataset using the cyclopamine accumulation profile as a predefined model for gene expression with the pattern‐matching algorithm Haystack. Refactoring candidate genes in Sf9 insect cells led to discovery of four enzymes that catalyze the first six steps in steroid alkaloid biosynthesis to produce verazine, a predicted precursor to cyclopamine. Three of the enzymes are cytochromes P450 while the fourth is a γ‐aminobutyrate transaminase; together they produce verazine from cholesterol. |
| Keywords: | verazine Veratrum californicum California corn lily Haystack steroid alkaloids cyclopamine |