Discovery of genes converting toxic tomato glycoalkaloids to edible esculeosides

Overview

This research addresses a key question in tomato fruit biology and food safety: how toxic, bitter steroidal glycoalkaloids (SGAs) are naturally converted into edible esculeosides during tomato ripening. While cultivated tomatoes are safe to eat, wild relatives and some breeding lines accumulate harmful SGAs. Only one gene in this conversion pathway (the C-23 hydroxylase GAME31) has been identified to date. The remaining enzymes that perform sterol acetylation, hydroxylation, and glycosylation, along with the transcription factors that regulate them, remain unknown. This project seeks to identify these missing genes, enabling future applications in tomato breeding, food safety, and metabolic engineering.

Technical specifications

Research approach:

• RNA sequencing (Illumina short-read and Oxford Nanopore long-read) of tomato fruit tissue (Micro-Tom cultivar) collected at eight developmental time points with five biological replicates
• Gene co-expression analysis using in-house Python code to identify genes whose expression correlates with GAME31 (Pearson correlation greater than 95%, p-value less than 0.01)
• Candidate gene prioritization through protein sequence homology searches against enzymes known to be involved in sterol metabolism
• Functional characterization of candidate genes encoding the three remaining enzymatic steps and the regulatory transcription factor(s)

Scientific rationale: Genes involved in related specialized metabolic pathways in tomato co-express because they are activated by shared transcription factors, such as the known regulator GAME9. Therefore, genes involved in the SGA-to-esculeoside conversion should display similar co-expression patterns, making transcriptomics a powerful discovery tool.

Technology readiness level

This is an early-stage fundamental research project (TRL 1–2). The first gene in the pathway (GAME31) has been identified, but the remaining enzymes and transcription factors are not yet characterized. The proposed RNA sequencing, co-expression analysis, and homology-based candidate prioritization represent the discovery and validation phases. Functional characterization of candidate genes would constitute the next stage of development toward potential applications in crop improvement and food safety.