Standard-flow targeted proteomics for quantification of enzymes in glucosinolate biosynthetic pathways

Abstract

Insights gained from quantitative proteomics studies show that the relationship between transcript and protein abundance is not generally linear. Quantitative measurement of both proteins and transcripts is required to reveal their actual relationship. Whereas quantitative analysis of transcripts is widely established, there is a need for easily accessible protein level technologies. Mass spectrometry-based proteomics is a promising solution, provided that it can be made accessible to a broader user group than it currently is. Developments in proteomics methodology show that relatively simple instrumentation allows for targeted proteomics which provides a reliable and accessible technology platform for protein quantification. I here describe the successful implementation of standard-flow targeted proteomics and its use to quantify 12 enzymes in the glucosinolate biosynthetic pathways in Arabidopsis thaliana. Enabled by targeted proteomics, I analyzed a series of transcription factor mutants and found that transcript and protein generally correlated well under these unchallenged conditions. When I treated wild-type plants with the phytohormone methyl-jasmonate, I observed uncoupling of metabolite, transcript and protein levels, suggesting post-transcriptional and/or -translational in addition to transcriptional regulation.