Elucidating steroid alkaloid biosynthesis in Veratrum californicum: production of verazine in Sf9 cells.
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.
Donald Danforth Plant Science Center, 975 North Warson Road, St. Louis, MO, 63132, USA.
Megan M
MM
Augustin
Donald Danforth Plant Science Center, 975 North Warson Road, St. Louis, MO, 63132, USA.|Monsanto Company, 700 Chesterfield Parkway West, St Louis, MO, 63017, USA.
Dan R
DR
Ruzicka
Donald Danforth Plant Science Center, 975 North Warson Road, St. Louis, MO, 63132, USA.|CSIR-Central Institute of Medicinal and Aromatic Plants, P.O. CIMAP, Lucknow, 226015, Uttar Pradesh, India.
Ashutosh K
AK
Shukla
Donald Danforth Plant Science Center, 975 North Warson Road, St. Louis, MO, 63132, USA.
Jörg M
JM
Augustin
Sequoia Sciences, 1912 Innerbelt Business Center Dr, St. Louis, MO, 63114, USA.
Courtney M
CM
Starks
Sequoia Sciences, 1912 Innerbelt Business Center Dr, St. Louis, MO, 63114, USA.
Mark
M
O'Neil-Johnson
Donald Danforth Plant Science Center, 975 North Warson Road, St. Louis, MO, 63132, USA.
Michael R
MR
McKain
Donald Danforth Plant Science Center, 975 North Warson Road, St. Louis, MO, 63132, USA.
Bradley S
BS
Evans
Botanic Gardens and Parks Authority, Kings Park and Botanic Garden, Fraser Avenue, West Perth 6005, WA, Australia.|School of Plant Biology, University of Western Australia, 35 Stirling Highway, Crawley, Perth, 6009, WA, Australia.
Matt D
MD
Barrett
Botanic Gardens and Parks Authority, Kings Park and Botanic Garden, Fraser Avenue, West Perth 6005, WA, Australia.|School of Plant Biology, University of Western Australia, 35 Stirling Highway, Crawley, Perth, 6009, WA, Australia.
Ann
A
Smithson
Department of Biological Sciences, University of Alberta, Edmonton, AB, Canada.|Department of Medicine, University of Alberta, Edmonton, AB T6G 2E9, Canada.|BGI-Shenzhen, Build 11, Beishan Industrial Zone, Yantian District, Shenzhen, 518083, China.
Gane Ka-Shu
GK
Wong
University of Alberta, 116 St and 85 Ave, Edmonton, AB T6G 2R3, Canada.
Michael K
MK
Deyholos
Division of Biological Sciences, 311 Bond Life Sciences Center, University of Missouri, Columbia, MO, 65211, USA.|Department of Plant and Microbial Biology, University of California, Berkeley, CA, 94720, USA.
Patrick P
PP
Edger
Division of Biological Sciences, 311 Bond Life Sciences Center, University of Missouri, Columbia, MO, 65211, USA.
J Chris
JC
Pires
Franklin College of Arts and Sciences, University of Georgia, Athens, GA, 30602, USA.
James H
JH
Leebens-Mack
Infinity Pharmaceuticals, 780 Memorial Drive, Cambridge, MA, 02139, USA.|Cellular Dynamics International, 525 Science Drive, Madison, WI, 53711, USA.
David A
DA
Mann
Donald Danforth Plant Science Center, 975 North Warson Road, St. Louis, MO, 63132, USA.
Toni M
TM
Kutchan
D000612:4-Aminobutyrate Transaminase / Q000235:genetics / Q000378:metabolism; D000465:Algorithms; D000818:Animals; D053898:Biosynthetic Pathways; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D004798:Enzymes / Q000235:genetics / Q000378:metabolism*; D020869:Gene Expression Profiling / Q000379:methods; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D017423:Sequence Analysis, RNA / Q000379:methods; D061987:Sf9 Cells; D059467:Transcriptome; D014703:Veratrum / Q000235:genetics* / Q000378:metabolism*; D014704:Veratrum Alkaloids / Q000378:metabolism*
D004798:Enzymes; D010940:Plant Proteins; D014704:Veratrum Alkaloids; C052979:verazine; D003577:Cytochrome P-450 Enzyme System; D000612:4-Aminobutyrate Transaminase; C000541:cyclopamine
California corn lily; Haystack; Veratrum californicum; cyclopamine; steroid alkaloids; verazine