The small subunit of snapdragon geranyl diphosphate synthase modifies the chain length specificity of tobacco geranylgeranyl diphosphate synthase in planta.
Geranyl diphosphate (GPP), the precursor of many monoterpene end products, is synthesized in plastids by a condensation of dimethylallyl diphosphate and isopentenyl diphosphate (IPP) in a reaction catalyzed by homodimeric or heterodimeric GPP synthase (GPPS). In the heterodimeric enzymes, a noncatalytic small subunit (GPPS.SSU) determines the product specificity of the catalytic large subunit, which may be either an active geranylgeranyl diphosphate synthase (GGPPS) or an inactive GGPPS-like protein. Here, we show that expression of snapdragon (Antirrhinum majus) GPPS.SSU in tobacco (Nicotiana tabacum) plants increased the total GPPS activity and monoterpene emission from leaves and flowers, indicating that the introduced catalytically inactive GPPS.SSU found endogenous large subunit partner(s) and formed an active snapdragon/tobacco GPPS in planta. Bimolecular fluorescence complementation and in vitro enzyme analysis of individual and hybrid proteins revealed that two of four GGPPS-like candidates from tobacco EST databases encode bona fide GGPPS that can interact with snapdragon GPPS.SSU and form a functional GPPS enzyme in plastids. The formation of chimeric GPPS in transgenic plants also resulted in leaf chlorosis, increased light sensitivity, and dwarfism due to decreased levels of chlorophylls, carotenoids, and gibberellins. In addition, these transgenic plants had reduced levels of sesquiterpene emission, suggesting that the export of isoprenoid intermediates from the plastids into the cytosol was decreased. These results provide genetic evidence that GPPS.SSU modifies the chain length specificity of phylogenetically distant GGPPS and can modulate IPP flux distribution between GPP and GGPP synthesis in planta.
Orlova
Irina
I
Department of Horticulture and Landscape Architecture, Purdue University, West Lafayette, Indiana 47907, USA.
Nagegowda
Dinesh A
DA
Kish
Christine M
CM
Gutensohn
Michael
M
Maeda
Hiroshi
H
Varbanova
Marina
M
Fridman
Eyal
E
Yamaguchi
Shinjiro
S
Hanada
Atsushi
A
Kamiya
Yuji
Y
Krichevsky
Alexander
A
Citovsky
Vitaly
V
Pichersky
Eran
E
Dudareva
Natalia
N
D032161:Antirrhinum / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D011756:Diphosphates / Q000378:metabolism; D004224:Diterpenes / Q000378:metabolism; D051231:Farnesyltranstransferase / Q000235:genetics / Q000378:metabolism*; D035264:Flowers / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D039821:Monoterpenes / Q000378:metabolism; D018515:Plant Leaves / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000201:enzymology / Q000235:genetics; D018749:RNA, Plant / Q000235:genetics; D012717:Sesquiterpenes / Q000378:metabolism*; D014026:Nicotiana / Q000201:enzymology* / Q000235:genetics
D011756:Diphosphates; D004224:Diterpenes; D039821:Monoterpenes; D018749:RNA, Plant; D012717:Sesquiterpenes; C511282:geranyl diphosphate; D051231:Farnesyltranstransferase