Speakers - 2026

Title: Revisiting ORCA3: A stress-inducible controller of camptothecin biosynthesis

Abstract

Camptothecin (CPT) is a monoterpene indole alkaloid and is one of the promising anticancer compounds. The genus Ophiorrhiza has emerged as a potential alternative source for CPT because a significant amount of camptothecin is present throughout the plant. To understand CPT biosynthesis, different in vitro tissues of O. rugosa var. decumbens were analyzed for their CPT content and relative expression of the candidate genes involved in biosynthesis. Of all the tissues, callus, rooted callus, and hairy root were selected, as callus, being the undifferentiated tissue, showed only traces of CPT. Rooted callus had three-fold higher CPT than callus with just slight differentiation and development of adventitious roots in it. Whereas hairy root exceeded the rooted callus by 10-fold. A comprehensive RNA-seq-based transcriptomic analysis was performed, providing a few potential candidate genes.

The present study focuses on understanding the role of ORCA3 (Octadecanoid–derivative Responsive Catharanthus AP2-domain protein 3) – a transcription factor as a master regulator of camptothecin biosynthesis in O rugosa. Seven sequences of ORCA3 were identified, among which ORCA3.2 showed a positive correlation with CPT. The size of the sequence was 591bp, translating to a protein of 196 amino acids with an expected molecular weight of 21.5 kDa. Transgenic O. rugosa lines overexpressing ORCA3.2 exhibited upregulation of several key CPT biosynthetic genes, including STR2, STR4, SLS2, SLS5, CPR3, LAMT, and 10-HGO, along with enhanced camptothecin content. To understand the regulation of ORCA3.2, the promoter of ORCA3.2 was identified and cloned upstream of GUS in pCAMBIA1301. Among various elicitors, ORCA3.2 showed the strongest activation by methyl jasmonate, followed by abscisic acid and gibberellic acid, indicating involvement of hormone and stress-mediated regulation. The study offers novel insights into ORCA3.2- mediated CPT biosynthesis regulation in O. rugosa, and may have potential applications in pathway engineering and elicitor-mediated enhancement of CPT content in the target plant.

The audience take away from presentation:

1. They can gain new insights into camptothecin biosynthesis that can support future studies on medicinal plants and facilitate the development of improved production strategies for pharmaceutically important metabolites.
2. Utilize promoter and elicitor analysis to investigate hormone-responsive regulation of biosynthetic pathways and identify conditions that can improve metabolite accumulation.
3. Understand a transferable research workflow integrating metabolite profiling, RNA-seq, gene expression analysis, transgenic validation, and promoter studies to identify key regulatory genes controlling specialized metabolite biosynthesis.
4. Expand research or teaching in plant biotechnology by using this study as a case example of functional genomics, transcriptional regulation, and pathway engineering in medicinal plants.