Studies of a biochemical factory: tomato trichome deep expressed sequence tag sequencing and proteomics.
Shotgun proteomics analysis allows hundreds of proteins to be identified and quantified from a single sample at relatively low cost. Extensive DNA sequence information is a prerequisite for shotgun proteomics, and it is ideal to have sequence for the organism being studied rather than from related species or accessions. While this requirement has limited the set of organisms that are candidates for this approach, next generation sequencing technologies make it feasible to obtain deep DNA sequence coverage from any organism. As part of our studies of specialized (secondary) metabolism in tomato (Solanum lycopersicum) trichomes, 454 sequencing of cDNA was combined with shotgun proteomics analyses to obtain in-depth profiles of genes and proteins expressed in leaf and stem glandular trichomes of 3-week-old plants. The expressed sequence tag and proteomics data sets combined with metabolite analysis led to the discovery and characterization of a sesquiterpene synthase that produces beta-caryophyllene and alpha-humulene from E,E-farnesyl diphosphate in trichomes of leaf but not of stem. This analysis demonstrates the utility of combining high-throughput cDNA sequencing with proteomics experiments in a target tissue. These data can be used for dissection of other biochemical processes in these specialized epidermal cells.
Schilmiller
Anthony L
AL
Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, Michigan 48824-1319, USA. schilmil@msu.edu
Miner
Dennis P
DP
Larson
Matthew
M
McDowell
Eric
E
Gang
David R
DR
Wilkerson
Curtis
C
Last
Robert L
RL
D053898:Biosynthetic Pathways; D002726:Chlorogenic Acid / Q000737:chemistry; D018076:DNA, Complementary / Q000235:genetics; D020224:Expressed Sequence Tags / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics; D018551:Solanum lycopersicum / Q000033:anatomy & histology* / Q000166:cytology / Q000201:enzymology / Q000235:genetics*; D055432:Metabolomics; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000166:cytology / Q000235:genetics; D018547:Plant Stems / Q000166:cytology / Q000235:genetics; D040901:Proteomics / Q000379:methods*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012431:Rutin / Q000096:biosynthesis / Q000737:chemistry; D017422:Sequence Analysis, DNA / Q000379:methods*; D013729:Terpenes / Q000378:metabolism
D018076:DNA, Complementary; D012333:RNA, Messenger; D013729:Terpenes; D002726:Chlorogenic Acid; D012431:Rutin