3rd Edition Of Plant Science and Molecular Biology World Conference 2026

Abstract

Trichomes are specialized epidermal structures that contribute to plant defense against environmental and biotic stresses. Although the genetic framework underlying unicellular trichome development has been extensively characterized, the regulatory mechanisms controlling multicellular trichome development remain insufficiently understood. Here, we identify a regulatory module involving Hair3 (H3) and Hair4 (H4), two C2H2-type zinc finger proteins, in tomato (Solanum lycopersicum). Functional analyses using CRISPR-Cas9-generated mutants revealed that single knockout of h3 or h4 plants exhibited no detectable trichome defects, whereas h3/h4 double (h3/h4-dko) mutants displayed pronounced alterations in trichome patterning. Specifically, the h3/h4-dko mutant showed reduced densities of glandular trichomes, but increased densities of non-glandular trichomes, accompanied by shorter long trichomes in leaves and stems. These results indicate that H3 and H4 function redundantly yet are essential for maintaining proper trichome composition and morphology. Biochemical assays demonstrated that H3 and H4 form both homodimers and heterodimers, supporting their cooperative function. Transcriptome and expression analyses further revealed that H3 and H4 regulate key genes associated with trichome development, including Woolly (Wo) and its downstream targets (Wox3b, MX1, H, and HD8). Although H3 and H4 did not directly bind to the Wo promoter, protein interaction assays showed that they physically associate with Wo, thereby enhancing the expression of Wo and Wox3b. Collectively, these findings establish H3 and H4 as central components of a regulatory network governing multicellular trichome development and provide new insights into the transcriptional mechanisms underlying epidermal differentiation in tomato. In addition, several differentially expressed genes identified in the transcriptome dataset suggest that H3 and H4 may regulate additional, yet-uncharacterized factors involved in epidermal differentiation and developmental signaling, highlighting the broader regulatory scope and future research for potential genetic resoureces.