# Dissecting the Molecular Mechanism of 10-HDA Biosynthesis: Role of Acyl-CoA Delta(11) Desaturase and Transcriptional Regulators in Honeybee Mandibular Glands

## TL;DR

This study identifies the genes and enzymes involved in producing 10-HDA, a key fatty acid in royal jelly, in honeybee glands.

## Contribution

The study provides the first complete molecular model of 10-HDA biosynthesis and identifies key regulatory genes in honeybee mandibular glands.

## Key findings

- A five-step biosynthetic pathway for 10-HDA was identified with conserved enzyme expression across two honeybee species.
- Knockdown of d11ds (LOC551527) reduced 10-HDA levels by over 50%, confirming its role in biosynthesis.
- Transcriptional regulators Kay and Drep-2 were identified as potential modulators of 10-HDA metabolism.

## Abstract

Honeybee workers utilize specialized mandibular glands (MGs) to produce 10-hydroxy-2-decenoic acid (10-HDA), a fatty acid unique and critical for royal jelly. Despite its nutritional and therapeutic significance, the molecular basis of 10-HDA biosynthesis remained unresolved. This study elucidated the production mechanism by analyzing gene expression patterns in honeybee glands across developmental stages and comparing these processes between two closely related species (Apis mellifera and A. cerana). We identified five interconnected metabolic steps governing 10-HDA synthesis, revealing conserved regulatory patterns in both species. MGs with elevated 10-HDA output exhibited enhanced activity of fatty acid-processing proteins while suppressing a key enzyme (encoded by d11ds, LOC551527) responsible for modifying fatty acid structures, reducing the 10-HDA levels by over 50%, directly linking this gene to biosynthesis. PPI network analysis further highlighted transcriptional regulators Kay and Drep-2 as potential modulators of 10-HDA metabolism. These findings establish the first complete model of 10-HDA production, demonstrating how the fatty acid metabolism of honeybee MGs adapts to worker roles.

10-Hydroxy-2-decenoic acid (10-HDA), a major fatty acid (FA) component of royal jelly, is synthesized in the mandibular glands (MGs) of worker honeybees. Despite its well-documented nutritional and therapeutic significance, the biosynthetic pathway and regulatory mechanisms of 10-HDA production remain largely unresolved. In this study, the molecular basis of 10-HDA biosynthesis and regulation in the MGs of newly emerged bees (NEBs), nurse bees (NBs), and forager bees (FBs) were investigated using RNA sequencing and weighted gene co-expression network analysis (WGCNA). A five-step biosynthetic pathway for 10-HDA was proposed, and cross-species analysis of Apis mellifera and A. cerana revealed the conserved expression patterns of 15 key enzymes involved. Functional validation via RNA interference (RNAi) demonstrated that knockdown of acyl-CoA Delta(11) desaturase (d11ds, LOC551527), a key enzyme in FA desaturation, led to a 50% reduction in 10-HDA levels. Protein–protein interaction (PPI) network analysis further identified transcriptional regulators Kay and Drep-2 as potential modulators of 10-HDA metabolism. This study provides the first comprehensive mechanistic model of 10-HDA biosynthesis in honeybee MGs and highlights the labor-specific regulation of FA metabolism. These findings offer promising genetic targets for improving the royal jelly quality through genetic technology.