Ascorbic acid modulation by ABI4 transcriptional repression of VTC2 in the salt tolerance of Arabidopsis.

BACKGROUND: Abscisic acid (ABA) plays an important role in plant abiotic stress responses, and ABA INSENSITIVE 4 (ABI4) is a pivotal transcription factor in the ABA signaling pathway. In Arabidopsis, ABI4 negatively regulates salt tolerance; however, the mechanism through which ABI4 regulates plant salt tolerance is poorly understood. Our previous study showed that ABI4 directly binds to the promoter of the VITAMIN C DEFECTIVE 2 (VTC2) gene, inhibiting the transcription of VTC2 and ascorbic acid (AsA) biosynthesis. RESULTS: In the present study, we found that treatment with exogenous AsA could alleviate salt stress sensitivity of ABI4-overexpressing transgenic plants. The decreased AsA content and increased reactive oxygen species (ROS) levels in ABI4-overexpressing seedlings under salt treatment indicated that AsA-promoted ROS scavenging was related to ABI4-mediated salt tolerance. Gene expression analysis showed that ABI4 was induced at the early stage of salt stress, giving rise to reduced VTC2 expression. Accordingly, the abundance of the VTC2 protein decreased under the same salt stress conditions, and was absent in the ABI4 loss-of-function mutants, suggesting that the transcriptional inhibition of ABI4 on VTC2 resulted in the attenuation of VTC2 function. In addition, other encoding genes in the AsA biosynthesis and recycling pathways showed different responses to salt stress, demonstrating that AsA homeostasis is complicated under salinity stress. CONCLUSIONS: This study elucidates the negative modulation of ABI4 in salt stress tolerance through the regulation of AsA biosynthesis and ROS accumulation in plants.

Ascorbic Acid Integrates the Antagonistic Modulation of Ethylene and Abscisic Acid in the Accumulation of Reactive Oxygen Species.

During plant growth and development, ethylene and abscisic acid (ABA) play important roles and exert synergistic or antagonistic effects on various biological processes, but the detailed mechanism underlying the interaction of the two phytohormones, especially in the regulation of the accumulation of reactive oxygen species (ROS), is largely unclear. Here, we report that ethylene inhibits but ABA promotes the accumulation of ROS in Arabidopsis (Arabidopsis thaliana) seedlings. Furthermore, changes in the biosynthesis of ascorbic acid (AsA) act as a key factor in integrating the interaction of ethylene and ABA in the regulation of ROS levels. We found that ethylene and ABA antagonistically regulate AsA biosynthesis via ETHYLENE-INSENSITIVE3 (EIN3) and ABA INSENSITIVE4 (ABI4), which are key factors in the ethylene and ABA signaling pathways, respectively. In addition, ABI4 is transcriptionally repressed by EIN3 in ethylene-regulated AsA biosynthesis. Via transcriptome analysis and molecular and genetic experiments, we identified VITAMIN C DEFECTIVE2as the direct target of ABI4 in the regulation of AsA biosynthesis and ROS accumulation. Thus, the EIN3-ABI4- VITAMIN C DEFECTIVE2 transcriptional cascade involves a mechanism by which ethylene and ABA antagonistically regulate AsA biosynthesis and ROS accumulation in response to complex environmental stimuli.

Deletion of clock gene Per2 exacerbates cholestatic liver injury and fibrosis in mice.

The Period 2 (Per2) gene is an important component of the circadian system and is thought to modulate many physiological and pathological processes in mammals. In the previous study, we have disclosed the protective role of Per2 against carbon tetrachloride induced liver injury and fibrosis. Here we further assess the effect of Per2 deficiency on cholestatic hepatic injury and fibrosis. Cholestasis was induced by bile duct ligation (BDL) for 10 days in wild-type (WT) and Per2(-/-) mice. Masson trichrome staining and analysis of α-SMA immunohistochemistry were performed to show the collagen accumulation and the HSC activation, respectively. The mRNA levels of fibrosis-related genes were monitored by quantitative real-time PCR. Following BDL, livers from Per2(-/-) mice exhibited markedly increased extent of bile infarct and extracellular matrix (ECM) deposition compared with WT mice. Furthermore, the expressions of fibrosis-related genes like TNF-α, TGF-ß1, Col1α1 and TIMP-1 were dramatically elevated in Per2(-/-) cholestatic liver. Our observations indicated that clock gene Per2 plays a protective role in mediating liver injury and fibrosis during cholestasis.

Clock gene mPer2 functions in diurnal variation of acetaminophen induced hepatotoxicity in mice.

The circadian clock gene Period2 (Per2) plays important roles in many physiologic and pathologic processes in mammals. In the previous study, we have reported the protective role of mPer2 against carbon tetrachloride induced hepatotoxicity. Here, we further explore the contribution of this gene to acetaminophen (APAP) induced liver injury in mice. It is reported that the hepatotoxicity induced by APAP exhibited a circadian rhythm in which the peak sensitive injection time is 20:00 while when the administration time becomes to 08:00, it caused markedly decreased liver damage. Thus, we injected APAP into wide type (WT) and mPer2 null mice at the dose of 300 mg/kg at both 08:00 and 20:00. Interestingly, the liver damage showed no significant difference between WT and mPer2 null mice when administered at 08:00, however, liver injury occurred in mPer2 null mice displayed less severe than WT at 20:00. In addition, Cyp1a2, one of the most important cytochrome P450 isoforms responsible for APAP bioactivation, decreased mRNA level at 20:00 in mPer2 null mice while its expression was not different in both strain mice at 08:00. Coincidently, the hepatic circadian rhythm expression of Per2 revealed that its mRNA level was weak at 08:00 but reached peak expression during 24 h at 20:00 in WT mice. Therefore, it is speculated that clock gene mPer2 may function in diurnal variation of APAP induced hepatotoxicity via modulating Cyp1a2 expression in mice.

Altered circadian rhythm of the clock genes in fibrotic livers induced by carbon tetrachloride.

Disruption in circadian rhythms either by mutation in mice or by shiftwork in people, is associated with an increased risk for the development of multiple organ diseases. In turn, organ disease may influence the function of clock genes and peripheral circadian systems. Here we showed that hepatic fibrosis induced by carbon tetrachloride in mice leads to alterations in the circadian rhythms of hepatic clock genes. Especially, we found an impaired daily Cry2 rhythm in the fibrotic livers, with markedly decreased levels during the day time while compared with control livers. Associatively, the expressions of two important clock-regulated genes peroxisome proliferator-activated receptor alpha and cytochrome P450 oxidoreductase lost circadian rhythm with significantly decreased levels during the light-dark (12/12h) cycle in fibrotic livers.