Metabolic engineering of tobacco plants using Carica papaya NAC transcription factors: a biotechnology tool to improve plant metabolite production.

Plant secondary metabolism includes the biosynthesis of a wide array of molecules with significant biological and biotechnological applications. However, the regulatory mechanisms governing secondary metabolite production remain largely elusive. Key master regulators of gene expression, such as transcription factors (TFs), are expected to play a central role in controlling the biosynthesis of these compounds. In this context, the present study addresses a critical knowledge gap by elucidating the regulatory role of Carica papaya NAC transcription factors (CpNACs) in reprogramming secondary metabolites in Nicotiana tabacum. Thus, this work advances our understanding of plant metabolic engineering by identifying molecular links between transcription factor regulation and metabolite accumulation. Methanolic extracts from transformed and non-transformed plants were analyzed by gas chromatography-mass spectrometry (GC-MS). The results showed altered accumulation of some specialized metabolites. Specifically, targeted metabolomics revealed increased accumulation of phytosterols, including campesterol, β-sitosterol, and stigmasterol. Non-targeted metabolomics further indicated extensive metabolic reprogramming, affecting pools of organic acids, steroids, and lipids. Furthermore, transformed seedlings showed elevated levels of phenylpropanoids (e.g., coumarin, sinapaldehyde), alkaloids (e.g., tropinone, allocryptopine), and derivatives of aromatic amino acids (e.g., tryptamine, phenylacetic acid). The increased abundance of key metabolic intermediates, such as acetate and acetyl-CoA, suggests a redirection of metabolic flux toward the biosynthesis of these specialized compounds. Our findings suggest that CpNAC transcription factors may regulate specialized metabolite biosynthetic pathways, potentially by modulating metabolic flux at specific enzymatic steps within these networks. This pioneering study proposes an alternative that could improve the performance and productivity of biotechnological processes for obtaining valuable compounds from plants, highlighting the potential of transcription factor-based approaches to manipulate complex plant metabolic networks.