Genome-wide analysis of the HbWOX gene family and HbWOX14-mediated enhancement of hairy root transformation in Hevea brasiliensis

Background The WOX gene family, a plant-specific group of essential transcription factors, plays important regulatory roles in stem cell maintenance, organogenesis, and hormone response cascades. For rubber trees, genotype-dependent limitations in tissue culture and low genetic transformation efficiency have long posed bottlenecks to their molecular breeding progress, where optimizing root regeneration systems is a critical breakthrough point. Results To address these challenges, we identified 17 HbWOX family members from the GT1 genome of rubber trees using bioinformatic approaches, and systematically analyzed their gene structures, conserved motifs, cis-acting elements, and tissue-specific expression profiles. Our findings revealed obvious copy-number diversification within the HbWOX family. Most members harbor a conserved homeodomain (HD) and share the specific motif "PEVSSRWNPTPEQLR", reflecting evolutionary conservation and functional specialization of this gene family in rubber trees. Notably, HbWOX14 exhibited root-specific and high-level expression, with its promoter region significantly enriched for auxin-responsive cis-elements. Functional verification showed that HbWOX14 responds rapidly to auxins such as IAA, and its expression pattern is tightly associated with root development processes. Using an Agrobacterium rhizogenes -mediated transformation system, overexpression of HbWOX14 increased the induction rate of rubber tree hairy roots from 13.3% to 66.7%, while simultaneously significantly improving root length, number, and thickness. Transcriptome and Gene Ontology (GO) enrichment analyses further demonstrated that HbWOX14 exerts its regulatory function by activating the auxin signaling pathway and modulating the expression of genes involved in cell division and differentiation. Additionally, it suppresses abscisic acid and jasmonic acid-mediated stress response pathways while coordinating flavonoid glycosylation-related secondary metabolic processes, collectively creating a favorable molecular environment for hairy root development. Conclusion This study is the first to systematically clarify the evolutionary characteristics and functional divergence of the HbWOX gene family in rubber trees, with a specific focus on verifying the key role of HbWOX14 in enhancing hairy root regeneration efficiency. These findings not only enrich our understanding of the plant WOX gene family's functional diversity but also provide crucial theoretical basis and practical technical support for accelerating rubber tree molecular breeding and improving its genetic transformation system.