Tongxinluo Reverses the Hypoxia-suppressed Claudin-9 in Cardiac Microvascular Endothelial Cells

Background： Claudin-5, claudin-9, and claudin-11 are expressed in endothelial cells to constitute tight junctions, and their deficiency may lead to hyperpermeability, which is the initiating process and pathological basis of cardiovascular disease.Although tongxinluo （TXL） has satisfactory antianginal effects, whether and how it modulates claudin-5, claudin-9, and claudin-1 1 in hypoxia-stimulated human cardiac microvascular endothelial cells （HCMECs） have not been reported.Methods： In this study, HCMECs were stimulated with CoCl2 to mimic hypoxia and treated with TXL.First, the messenger RNA （mRNA） expression of claudin-5, claudin-9, and claudin-l 1 was confirmed.Then, the protein content and distribution of claudin-9, as well as cell morphological changes were evaluated after TXL treatment.Furthermore, the distribution and content histone H3K9 acetylation （H3K9ac） in the claudin-9 gene promoter, which guarantees transcriptional activation, were examined to explore the underlying mechanism, by which TXL up-regulates claudin-9 in hypoxia-stimulated HCMECs.Results： We found that hypoxia-suppressed claudin-9 gene expression in HCMECs （F=7.244;P =0.011） and the hypoxia-suppressed claudin-9 could be reversed by TXL （F=61.911;P =0.000）, which was verified by its protein content changes （F=29.142;P =0.000）.Moreover, high-dose TXL promoted the cytomembrane localization of claudin-9 in hypoxia-stimulated HCMECs, with attenuation of cell injury.Furthermore, high-dose TXL elevated the hypoxia-inhibited H3K9ac in the claudin-9 gene promoter （F=37.766;P =0.000）, activating claudin-9 transcription.Conclusions： The results manifested that TXL reversed the hypoxia-suppressed claudin-9 by elevating H3K9ac in its gene promoter, playing protective roles in HCMECs.

WHIRLY1 recruits the histone deacetylase HDA15 repressing leaf senescence and flowering in Arabidopsis

Leaf senescence is controlled by a complex regulatory network in which robustness is ensured by the activity of transcription factors and epigenetic regulators.However,how these coordinate the process of leaf senescence remains poorly understood.We found that WHIRLY1 interacts with Histone Deacetylase(HDA)15,a Reduced Potassium Dependence3(RPD3)/HDA1-type HDA,by using green fluorescent protein-nanotrap-mass spectrum assays.The development-dependent interaction between WHIRLY1 and HDA15 was further confirmed by bimolecular fluorescence complementation assays and co-immunoprecipitation assays in Arabidopsis.Multi-omics genome-wide transcriptome and H3K9 acetylome enrichment analysis showed that HDA15 delays leaf senescence and flowering by repressing the expression of the positive regulators of leaf senescence and flowering,such as LOX2 and LARP1 C,and reducing H3K9 ac levels at these loci;WHIRLY1 and HDA15 co-target to the region near the transcription start site of a subset of nutrient recycling-related genes(e.g.,Glutathione S-transferases 10,non-coding RNA,and photosystem II protein D1 synthesizer attenuator PDIL1-2),as well as WRKY53 and ELF4,and co-repress their expression by removing H3K9 acetylation.Our study revealed a key transcription regulatory node of nutrient recycling and senescence-associated genes involved in leaf senescence and flowering via the recruitment of HDA15 by the single-stranded DNA/RNA-binding protein WHIRLY1.