# Identification of Central Regulatory Hubs in Pupal Diapause of Helicoverpa armigera Using Weighted Gene Co-Expression Network Analysis and Multiscale Embedded Network Analysis

## TL;DR

This study identifies key genes involved in the diapause process of Helicoverpa armigera, a pest insect, using advanced network analysis techniques.

## Contribution

The study introduces a systems-level view of diapause regulation and identifies novel candidate genes for pest control.

## Key findings

- Diapause involves extensive gene expression changes, particularly in IgSF CAM and nucleocytoplasmic transport pathways.
- Three candidate regulatory genes (DDX5, PLK4, TAF5L) were identified through network analysis.
- Differential gene expression increased over time, with persistent regulation in mitochondrial metabolism and hormone signaling.

## Abstract

Diapause is a programmed dormancy that helps insects survive unfavorable seasons. The cotton bollworm, Helicoverpa armigera, uses pupal diapause to overwinter, making it a widespread agricultural pest. Understanding how diapause is controlled at the molecular level could lead to new ways to manage this pest. In this study, we conducted a comprehensive, multi-faceted transcriptomic investigation, combining differential expression analysis, WGCNA, and MEGENA to progress from gene lists toward functional modules and central regulatory hubs. We found that diapause involves extensive changes in gene expression, especially in pathways related to IgSF CAM signaling and nucleocytoplasmic transport pathways, which have not been previously linked to insect diapause. Using gene network analysis, we identified three candidate genes, DDX5, PLK4, and TAF5L. These findings provide new insights into the genetic control of diapause and highlight potential targets for disrupting this process in pest populations.

Diapause is a vital overwintering strategy for many insects, yet its comprehensive molecular architecture remains elusive. In the polyphagous pest Helicoverpa armigera, facultative pupal diapause is key to its ecological success. To elucidate the complex diapause regulatory network, we conducted a transcriptomic analysis of diapause (DP) versus non-diapause (NP) pupal brains across early pupal development (days 2, 5, and 10). Integrated analyses, including differential expression, persistent gene identification, weighted gene co-expression network analysis (WGCNA), and multiscale embedded network analysis (MEGENA), were employed to define core regulatory modules and hubs. The number of differentially expressed genes (DEGs) increased over time, with 1781 genes persistently regulated across all time points, enriched in mitochondrial metabolism, hormone signaling, and chromatin remodeling. WGCNA revealed a diapause-associated module (red) linked to RNA processing/transcription and a development-associated module (blue) enriched for translation and mitochondrial metabolism. Network analyses pinpointed three central hub genes: DDX5 and PLK4 (downregulated in diapause, upregulated upon 20E treatment) and TAF5L (upregulated in diapause, downregulated after 20E). This study provides a systems-level view of the transcriptional landscape governing pupal diapause in H. armigera and identifies novel candidate regulators for future functional studies.