# Knocking Down miR172f in the Hairy Roots of Grass Pea Increases β-ODAP Content and Induces Global Transcriptomic Reprogramming

## TL;DR

Knocking down miR172f in grass pea hairy roots increases β-ODAP levels and changes gene activity, offering new insights into its biosynthesis.

## Contribution

miR172f is identified as a novel regulator of β-ODAP biosynthesis by targeting LsBAHD3 in grass pea.

## Key findings

- Knockdown of miR172f in L. sativus hairy roots increased β-ODAP content.
- miR172f targets LsBAHD3, a β-ODAP synthetase, to regulate its biosynthesis.
- Transcriptomic changes linked to protein ubiquitination and amino acid metabolism were observed.

## Abstract

Background: There is an abundance of the neuroactive β-N-oxalyl-L-α,β-diaminopropionic acid (β-ODAP) in grass pea (Lathyrus sativus), pea (Pisum sativum), and several Chinese traditional herbs such as Panax notoginseng. It is well known for its dose- and context-dependent effects on its toxicological characteristics (inducing neurodegenerative neurolathyrism upon excessive consumption) or for its pharmacological effects (including neuroprotection and wound healing). Therefore, reducing β-ODAP levels improves the safety profile of β-ODAP-containing species for utilization, whereas increasing them facilitates their isolation and purification. LsBAHD3 acyltransferase, named after the first letter of BEAT benzylalcohol O-acetyltransferase (BEAT), anthocyanin O-hydroxycinnamoyltransferase (AHCT), anthranilate N-hydroxycinnamoyl/benzoyltransferase (HCBT), and deacetylvindoline 4-Oacetyltransferase (DAT), was proven to be β-ODAP synthetase. Methods: In this report, the interaction of miR172f with LsBAHD3 was investigated through bioinformatic analysis and transient co-expression assays in Nicotiana benthamiana. Functions of miR172f in β-ODAP biosynthesis were also investigated through knockdown in the hairy roots of L. sativus and via transcriptomic analysis. Results: The results suggest that the knockdown of miR172f in hairy roots of L. sativus increased β-ODAP content via targets to LsBAHD3. In this process, protein ubiquitination, cysteine and methionine metabolism, enzyme regulator activity, and so on were associated with β-ODAP biosynthesis. Conclusions: These results identify miR172f as a novel regulator of β-ODAP biosynthesis through targeting of LsBAHD3, offering new insight into the gene expression of β-ODAP synthetase and the genetic network governing β-ODAP biosynthesis in L. sativus.