# Genome-Wide Characterization of DrRALF Genes in Yam (Dioscorea rotundata) Reveals Their Potential Roles in Tuber Expansion and the Gibberellin Response

## TL;DR

This study explores the role of RALF genes in yam tuber growth and their connection to gibberellin signaling, offering new insights for improving yam production.

## Contribution

The study identifies specific DrRALF genes involved in yam tuber expansion and their interaction with gibberellin signaling.

## Key findings

- Seven DrRALF genes were identified in the yam genome, with some specifically expressed in roots and flowers.
- DrRALF2, DrRALF4, DrRALF5, and DrRALF6 are regulated by gibberellin and its biosynthesis inhibitor.
- DrRALF6 interacts with the GA-signaling protein DrDELLA1, linking it to tuber expansion.

## Abstract

Yam (Dioscorea spp.) provides various nutritional and medicinal benefits, including a high starch content, dietary fiber, essential micronutrients, and bioactive compounds. The molecular mechanisms underlying tuber expansion have not yet been clarified. Rapid alkalinization factor (RALF) genes, which mediate various processes in plants, are thought to contribute to the regulation of tuber growth; however, their role in yam development, especially in gibberellin (GA)-mediated processes, remains unclear. Here, we characterized seven DrRALF genes in the yam genome. Analysis of gene duplication demonstrated that the expansion of DrRALF genes was primarily driven by whole-genome duplication or segmental duplication. Phylogenetic analysis revealed that DrRALF genes were concentrated in specific clusters, indicating that their functions are relatively conserved. DrRALF5 was specifically expressed in the roots, and DrRALF2, DrRALF3, DrRALF4, and DrRALF6 were highly expressed in flowers. DrRALF1, DrRALF2, DrRALF3, DrRALF4, DrRALF5, and DrRALF6 were shown to play a role in tuber expansion. Subsequent qRT-PCR validation of four selected DrRALF genes confirmed the regulation of DrRALF2, DrRALF4, DrRALF5, and DrRALF6 by GA and PP333 (paclobutrazol, a GA biosynthesis inhibitor). Yeast one-hybrid assays further showed that the DrRALF6 promoter region interacted with the GA-signaling protein, DrDELLA1. Our findings provide novel insights into the regulatory network controlling yam tuber expansion, especially through the interaction between DrRALF6 and GA signaling pathways. Our results clarify the molecular mechanisms involved in tuber growth and propose a promising strategy for improving yam production through genetic manipulation of the GA-RALF signaling pathway.