Effect of developmental dynamics on WRKY expression in barley with varying phenologies and trichome micromorphologies

Understanding how developmental traits and epidermal structures contribute to stress tolerance is essential for improving crop resilience. In this study, four spring barley genotypes differing in phenology and epidermal features—CamBW1, CamWa1, LubBW1, and LubWa1—were evaluated under combined abiotic (mild and severe drought) and biotic (Fusarium infection) stress, with growth regulators application. The first phase of the study focused on assessing the phenological diversity of the examined barley genotypes in order to characterize the role of earliness in response to combined abiotic and biotic stresses. Phenological analysis confirmed distinct developmental patterns between early- and late-heading genotypes, supporting the presence of a “drought escape” strategy in early-flowering lines. The expression of the key photoperiod gene HvPRR37 further reflected these phenological differences. Through a comprehensive screening of drought responses and Fusarium infection severity, we were able to determine the performance and stress tolerance of the studied genotypes, and subsequently explore the expression patterns of key transcription factors (WRKY). Scanning electron microscopy revealed clear variability in trichome micromorphology between genotypes: LubBW1 developed numerous small trichomes, whereas LubWa1 produced fewer but larger structures. These differences may underlie distinct responses to Fusarium infection, as LubBW1 trichomes appeared to trap fungal conidia more effectively. Photosynthetic performance analyses indicated that combined stress strongly affected photosystem II and I efficiency, with the most severe inhibition observed under drought + Fusarium + gibberellin application treatment. However, mild drought induced only moderate changes, suggesting adaptive physiological regulation. Gene expression profiling of WRKY34, WRKY51, and WRKY70 demonstrated genotype-specific and stress-intensity-dependent regulation. WRKY51 and WRKY70 responded rapidly to drought and pathogen stress, while WRKY34 expression appeared developmentally regulated. These results highlight the complex interplay between phenology, trichome micromorphology, photosynthetic efficiency, and transcriptional regulation in shaping barley responses to combined stresses, underscoring the developmental stage as a critical determinant of resistance activation.