HsfA1a upregulates melatonin biosynthesis to confer cadmium tolerance in tomato plants.

Melatonin regulates broad aspects of plant responses to various biotic and abiotic stresses, but the upstream regulation of melatonin biosynthesis by these stresses remains largely unknown. Herein, we demonstrate that transcription factor heat-shock factor A1a (HsfA1a) conferred cadmium (Cd) tolerance to tomato plants, in part through its positive role in inducing melatonin biosynthesis under Cd stress. Analysis of leaf phenotype, chlorophyll content, and photosynthetic efficiency revealed that silencing of the HsfA1a gene decreased Cd tolerance, whereas its overexpression enhanced plant tolerance to Cd. HsfA1a-silenced plants exhibited reduced melatonin levels, and HsfA1a overexpression stimulated melatonin accumulation and the expression of the melatonin biosynthetic gene caffeic acid O-methyltransferase 1 (COMT1) under Cd stress. Both an in vitro electrophoretic mobility shift assay and in vivo chromatin immunoprecipitation coupled with qPCR analysis revealed that HsfA1a binds to the COMT1 gene promoter. Meanwhile, Cd stress induced the expression of heat-shock proteins (HSPs), which was compromised in HsfA1a-silenced plants and more robustly induced in HsfA1a-overexpressing plants under Cd stress. COMT1 silencing reduced HsfA1a-induced Cd tolerance and melatonin accumulation in HsfA1a-overexpressing plants. Additionally, the HsfA1a-induced expression of HSPs was partially compromised in COMT1-silenced wild-type or HsfA1a-overexpressing plants under Cd stress. These results demonstrate that HsfA1a confers Cd tolerance by activating transcription of the COMT1 gene and inducing accumulation of melatonin that partially upregulates expression of HSPs.

Melatonin enhances thermotolerance by promoting cellular protein protection in tomato plants.

Melatonin is a pleiotropic signaling molecule that provides physiological protection against diverse environmental stresses in plants. Nonetheless, the mechanisms for melatonin-mediated thermotolerance remain largely unknown. Here, we report that endogenous melatonin levels increased with a rise in ambient temperature and that peaked at 40°C. Foliar pretreatment with an optimal dose of melatonin (10 µmol/L) or the overexpression of N-acetylserotonin methyltransferase (ASMT) gene effectively ameliorated heat-induced photoinhibition and electrolyte leakage in tomato plants. Both exogenous melatonin treatment and endogenous melatonin manipulation by overexpression of ASMT decreased the levels of insoluble and ubiquitinated proteins, but enhanced the expression of heat-shock proteins (HSPs) to refold denatured and unfolded proteins under heat stress. Meanwhile, melatonin also induced expression of several ATG genes and formation of autophagosomes to degrade aggregated proteins under the same stress. Proteomic profile analyses revealed that protein aggregates for a large number of biological processes accumulated in wild-type plants. However, exogenous melatonin treatment or overexpression of ASMT reduced the accumulation of aggregated proteins. Aggregation responsive proteins such as HSP70 and Rubisco activase were preferentially accumulated and ubiquitinated in wild-type plants under heat stress, while melatonin mitigated heat stress-induced accumulation and ubiquitination of aggregated proteins. These results suggest that melatonin promotes cellular protein protection through induction of HSPs and autophagy to refold or degrade denatured proteins under heat stress in tomato plants.

[Expression of tissue factor induced by IL-6 in HUVEC].

OBJECTIVE: To investigate the effects of IL-6 at the expression of tissue factor (TF) in human vein endothelial cells(HUVECs). METHODS: HUVECs were incubated with IL-6 at the concentration of 0.5 ng/mL. Cell viability was measured by CCK-8 assay. The TF mRNA was detected by reverse transcript-polymerase chain reaction(RT-PCR) method. RESULTS: When HUVECs were exposed to IL-6 (0.5 ng/mL) within a period of 72 h, their viability did not decrease in comparison with the control; there was no statistical difference between the two groups. After the HUVECs were exposed to IL-6 (0.5 ng/mL) for 6 h, the TF mRNA level increased, and it reached the peak at 12 h; then it began to decline. The expression of TF mRNA induced by IL-6 was evidently detected from 6 h to 48 h. After the HUVECs were treated by IL-6 over 72 h, the expression of TF mRNA was no longer detected in HUVECs. CONCLUSION: IL-6 at the concentration of 0.5 ng/mL did not exert direct effect on cell viability. The increase of TF mRNA expression in HUVECs induced by IL-6 could play an important role in the modulation of blood coagulation disorder and in the mechanism related to coagulation system changes during

[Effects of Porphyromonas gingivalis with different fimA genotypes on vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 production by human umbilical vein endothelial cells].

OBJECTIVE: To investigate the effect of Porphyromonas gingivalis (Pg) with different fimA genotypes on vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) production by human umbilical vein endothelial cells (HUVEC). METHODS: In the present study, PgATCC33277 (type I fimA genotype), WCSP 115 (type II fimA genotype), W83 (type IV fimA genotype), and Escherichia coli-lipopolysaccharide (Ec-LPS) were designed as experimental group 1, 2, 3, and positive control group, respectively, to stimulate HUVEC, and the un-stimulated HUVEC were analyzed as negative control group. The three strains of Pg were cultured anaerobically in standard condition, and then the Pg cells and Ec-LPS were co-cultured with HUVEC for 2, 6, and 24 h, respectively. The amount of ICAM-1 and VCAM-1 produced by HUVEC was detected with flow cytometry (FCM). The expression of ICAM-1 and VCAM-1 by HUVEC were assayed with confocal laser scanning microscope (CLSM). RESULTS: The expression of ICAM-1 on the surface of HUVEC were intensified after infected by Pg with I, II, and IV fimA genotypes (P < 0.05). The amounts of ICAM-1 were 60.27 ± 5.43, 80.81 ± 1.44, and 85.94 ± 2.56 for Pg with type I fimA genotype, 86.69 ± 8.81, 90.19 ± 0.00, and 96.18 ± 0.48 for Pg with type II fimA genotype, 59.66 ± 0.40, 85.79 ± 4.86, and 96.04 ± 2.07 for Pg with type IV fimA genotype at 2, 6 and 24 h after infection, respectively. The up-regulation effects caused by Pg with type II and IV fimA genotypes were stronger than those caused by Pg with type I fimA genotype at different time points except at 2 h (P < 0.05). Under the present experimental condition, infected by Pg with type I, II and IV fimA genotypes stimulated low expression of VCAM-1 by HUVEC, it showed no significant differences among all the groups (P > 0.05). Expression of ICAM-1 and VCAM-1 in Pg infected HUVEC were confirmed by CLSM. Infection of HUVEC with Pg resulted in more fluorescence staining of ICAM-1 and VCAM-1 compared with that in uninfected HUVEC cultures. CONCLUSIONS: The virulence and pathogenicity of Pg is associated with its fimA genotypes, Pg with type II and IV fimA genes possess stronger ability to stimulate HUVEC to up-regulate the expression of cell adhesion molecules, which may lead to disorders in vascular endothelial function.