The Dof (DNA binding with one finger) transcription factor family is a plant-specific group of transcription factors that play critical roles in plant growth and development, stress response, and the regulation of secondary metabolism. Prunella vulgaris (P. vulgaris) has attracted considerable attention due to its medicinal value, with rosmarinic acid being one of its key bioactive components. However, the systematic identification of the Dof transcription factor family in P. vulgaris and its regulatory role in rosmarinic acid biosynthesis remains poorly understood. In this study, based on the whole-genome data of P. vulgaris, we identified 48 Dof transcription factor genes distributed across 14 chromosomes using bioinformatics approaches. Physicochemical analysis revealed that the encoded proteins have molecular weights ranging from 15,482.44 to 55,875.53 Da, amino acid lengths between 142 and 509, and theoretical isoelectric points from 4.84 to 10.2. All proteins were predicted to be hydrophilic and localized in the nucleus. Phylogenetic analysis classified them into four subfamilies, and multiple sequence alignment confirmed that all members contain a conserved C2-C2-type zinc finger domain. Analysis of cis-regulatory elements in the promoter regions identified numerous elements related to light responsiveness, hormone response, and development. Transcriptomic expression profiling demonstrated distinct tissue-specific expression patterns of Dof genes, with some showing high expression in spikes and seeds. Correlation analysis between gene expression and rosmarinic acid content identified three candidate genes potentially involved in the regulation of rosmarinic acid biosynthesis, which were further validated by RT-qPCR. Moreover, protein–protein interaction network predictions indicated 242 interactions among 23 Dof proteins. This study provides the first systematic identification of the Dof transcription factor family in P. vulgaris, offering important insights into the transcriptional regulation of rosmarinic acid biosynthesis and presenting potential genetic targets for enhancing rosmarinic acid production through genetic engineering.

To overcome natural skin barrier, deliver ellagic acid (EA) to the dermis, and promote its anti-ageing efficacy, oligomeric hyaluronic acid (HA) modified EA-loaded liposomes (EA-HA-L) were constructed via self-synthesized different molecular weights of HA linked cholesterol (HA-Chol), and then the effect of HA molecular weight on the skin permeability of EA was explored to clarify the optimal molecular weight of HA with best transdermal delivery effectiveness. Finally, a series of in vitro and in vivo experiments were conducted to survey the transdermal mechanism, skin irritation, antioxidant, anti-photo ageing and antiwrinkle effects of EA-loaded liposomes modified with the optimal molecular weight of HA. The results showed that EA-HA-L had less than 200 nm particle size and high encapsulation efficiency. Among them, 5 kDa of oligomeric HA-modified liposomes (EA-HA5k-L) maximized the skin penetration and retention of EA and promoted the distribution width of EA in the skin far beyond the thickness of the epidermal layer, indicating its good ability to deliver EA to the dermis. EA-HA5k-L displayed uniformly sized nanosphere morphology and slow-release behavior in neutral and acidic environments that simulated skin. The transdermal mechanism of EA-HA5k-L was proven to be related to the loosening of the stratum corneum, reduction of calcium adhesion proteins, and recognition of CD44 receptor. EA-HA5k-L had no irritant effect on the chicken embryo chorioallantoic membrane, with an irritant index close to 0.9% NaCl. EA-HA5k-L not only improved the clearance rate of EA on DPPH and hydroxyl radicals but also elevated its inhibition effect on elastase. Significantly, compared to free EA and EA-loaded liposomes without oligomeric HA modification (EA-L), EA-HA5k-L significantly increased the cellular uptake of EA through receptor-mediated endocytosis, and effectively blocked the increase in metalloproteinase-1 (MMP-1) content and decrease in type I collagen content induced by UVB in human dermal fibroblasts (HDFs), demonstrating better anti-photo ageing effectiveness. Moreover, EA-HA5k-L upregulated the relative expression of the elastin gene and three types of type I collagen gene (col1a1a, col1a1b, and col1a2) in zebrafish, and its expression promotion rates in col1a1b, col1a2 and elastin were remarkably higher than those of free EA, EA-L, and acetyl hexapeptide-8 as positive control. Conclusively, EA-HA5k-L ameliorated the anti-ageing effectiveness of EA due to the successful transdermal delivery and efficient cellular uptake, and 5 kDa of oligomeric HA-modified liposomes may be a promising transdermal delivery carrier to overcome skin barrier and upgrade the application prospects of EA in anti-skin ageing.

Introduction: Selaginella species hold a traditional place in medicine and cosmetics, but their potential to extend lifespan and the underlying bioactive compounds remain inadequately investigated. This study aims to systematically evaluate the anti-aging properties of diverse Selaginella extracts and to identify the key bioactive components and mechanisms involved. Methods: We collected 23 Selaginella samples from 13 different provinces across China to assess their geographical influence. Two representative methanol extracts, S4 (high in amentoflavone) and S16 (low in amentoflavone), were selected for in-depth evaluation using the Caenorhabditis elegans model. We employed lifespan assays, stress resistance tests, and comparative transcriptomics to analyze the effects on longevity, and pathway modulation. Results: Both S4 and S16 extracts significantly extended the lifespan of C. elegans under normal conditions and modulated conserved longevity pathways, including MAPK and FOXO signaling, with daf-16 and egl-8 emerging as key hub genes. Amentoflavone was identified and validated as a critical bioactive component, which alone extended lifespan by 63.81% and enhanced stress resistance. Mechanistically, amentoflavone promoted the nuclear translocation of DAF-16 and up-regulated the expression of antioxidant genes (e.g., sod-3 , gst-3/4 , hsp-16.48/12.6 ), leading to a significant reduction in intracellular ROS levels. Discussion: Our findings demonstrate that Selaginella extract and its key component, amentoflavone, delay aging primarily by activating the DAF-16/FOXO transcription factor and bolstering the antioxidant defense system. This study not only highlights amentoflavone as a major contributor to the lifespan-extending effects of Selaginella but also underscores the potential of these natural compounds as promising agents for healthy aging.