Membrane-Localized Orientation of NONPHOTOTROPIC HYPOCOTYL3 Affects the Necessity of Its Phosphorylation for Phototropism.

NONPHOTOTROPIC HYPOCOTYL3 (NPH3) is a key regulator of hypocotyl phototropism under both low- and high-intensity blue light (LBL/HBL), mediating phototropin1 (phot1) and phot2 signaling. NPH3 undergoes dephosphorylation and is released from the plasma membrane (PM) upon blue light irradiation. However, how its phosphorylation status and PM localization mediate phot1 and phot2 signaling in Arabidopsis (Arabidopsis thaliana) remains elusive. In this study, we found that fusing NPH3 with GFP at its C terminus (N3G) impaired its release from the PM, a defect exacerbated by a phosphorylation-deficient mutation, resulting in a dephosphorylated NPH3-GFP (N3AG). Unlike N3G, transgenic lines expressing N3AG exhibited defective hypocotyl phototropism under HBL, which could be rescued by myristoylation at the N-terminus of N3AG (mN3AG), indicating that NPH3 phosphorylation is not essential for HBL-induced phototropic responses when it is artificially anchored at the PM via its N terminus. Furthermore, genetic analysis revealed that N3AG anchored to the PM by its N terminus (as in mN3AG) only rescues phot1-mediated HBL responses, which require RPT2. However, N3AG failed to regulate phot2-mediated HBL signaling, regardless of its PM orientation. Taken together, our results revealed that NPH3 phosphorylation is essential for phot2-mediated hypocotyl phototropism under HBL, but is not required for phot1-mediated HBL signaling when the NPH3 N terminus is PM-anchored.

Phototropin 1 Mediates High-Intensity Blue Light-Induced Chloroplast Accumulation Response in a Root Phototropism 2-Dependent Manner in Arabidopsis phot2 Mutant Plants.

Phototropins, namely, phototropin 1 (phot1) and phototropin 2 (phot2), mediate chloroplast movement to maximize photosynthetic efficiency and prevent photodamage in plants. Phot1 primarily functions in chloroplast accumulation process, whereas phot2 mediates both chloroplast avoidance and accumulation responses. The avoidance response of phot2-mediated chloroplasts under high-intensity blue light (HBL) limited the understanding of the function of phot1 in the chloroplast accumulation process at the HBL condition. In this study, we showed that the phot2 mutant exhibits a chloroplast accumulation response under HBL, which is defective when the root phototropism 2 (RPT2) gene is mutated in the phot2 background, mimicking the phenotype of the phot1 phot2 double mutant. A further analysis revealed that the expression of RPT2 was induced by HBL and the overexpression of RPT2 could partially enhance the chloroplast accumulation response under HBL. These results confirmed that RPT2 also participates in regulating the phot1-mediated chloroplast accumulation response under HBL. In contrast, RPT2 functions redundantly with neural retina leucine zipper (NRL) protein for chloroplast movement 1 (NCH1) under low-light irradiation. In addition, no chloroplast accumulation response was detected in the phot2 jac1 double mutant under HBL, which has been previously observed in phot2 rpt2 and phot1 phot2 double mutants. Taken together, our results indicated that phot1 mediates the HBL-induced chloroplast accumulation response in an RPT2-dependent manner and is also regulated by j-domain protein required for chloroplast accumulation response 1 (JAC1).

Protective effects of upregulated HO-1 gene against the apoptosis of human retinal pigment epithelial cells in vitro.

AIM: To investigate the protective effect of heme oxygenase-1 (HO-1) against H2O2-induced apoptosis in human ARPE-19 cells. METHODS: The lentiviral vector expressing HO-1 was prepared and transfected into apoptotic ARPE-19 cells induced by H2O2. Functional experiments including cell counting kit-8 (CCK-8) assay, flow cytometry (FCM) and mitochondrial membrane potential assay were conducted. RESULTS: The ultrastructure of ARPE-19 cells was observed using transmission electron microscope (TEM). It was found that exogenous HO-1 significantly ameliorated H2O2-induced loss of cell viability, apoptosis and intracellular levels of reactive oxygen species (ROS) in ARPE-19 cells. The overexpression of HO-1 facilitated the transfer of nuclear factor erythroid-2-related factor 2 (Nrf2) from cytoplasm to nucleus, which in turn upregualted expressions HO-1 and B-cell lymphoma-2 (Bcl-2). Furthermore, HO-1 upregulation further inhibited H2O2-induced release of cysteinyl aspartate specific proteinase-3 (caspase-3). CONCLUSION: Exogenous HO-1 protect ARPE-19 cells against H2O2-induced oxidative stress by regulating the expressions of Nrf2, HO-1, Bcl-2, and caspase-3.

[Comparison of the block characteristics of levobupivacaine vs bupivacaine for unilateral spinal block].

OBJECTIVE: To compare the block characteristics of levobupivacaine and bupivacaine for hypobaric unilateral spinal block. METHODS: Sixty patients undergoing hypobaric unilateral spinal block for lower extremity operations were randomized into 2 equal groups. In one group the patients were asked to lie on the uncompromised side for slow injection of hypobaric bupivacaine solution (0.75% 6 mg, diluted to 2 ml with distilled water) at the rate of 1 ml/15 s through a 27-gauge needle. The patients maintained their position for 15 min before shifting to a supine position. In the other group levobupivacaine was used instead. RESULTS: Statistical study showed that the block characteristics were of no significant difference between the 2 groups (P>0.05) except that the duration of motor block was shorter in levobupivacaine group than in bupivacaine group (P<0.05). Blood pressure or heart rate showed no significant difference between the two groups during operation. CONCLUSION: The block characteristics of hypobaric unilateral spinal block with levobupivacaine or bupivacaine are similar and their effect on circulatory function is slight. Levobupivacaine provides an option in case bupivacaine is not available.