PI3K is involved in L‐selectin‐ and PSGL‐1‐mediated neutrophil rolling on E‐selectin via F‐actin redistribution and assembly

L‐selectin and P‐selectin glycoprotein ligand‐1 (PSGL‐1) are adhesion molecules that play critical roles in neutrophil rolling during inflammation and lymphocyte homing. On the other hand they also function as signaling receptors to induce cytoskeleton changes. The present study is to investigate the signaling kinases responsible for the F‐actin changes mediated by L‐selectin and PSGL‐1 during neutrophil rolling on E‐selectin. Western blot analysis demonstrated that PI3K activation, peaking within 5 min, was induced by ligation of L‐selectin and PSGL‐1 with E‐selectin, and that Vav1 (the pivotal downstream effector of PI3K signaling pathway involved in cytoskeleton regulation) was recruited to the membrane and tyrosine‐phosphorylated, depending on PI3K. Furthermore, the F‐actin redistribution and assembly mediated by ligation with E‐selectin were blocked by LY294002, a PI3K specific inhibitor. Additional experiments showed that PI3K activity was involved in neutrophil rolling on E‐selectin. However, Syk/Zap70, the well‐known upstream kinase of PI3K, was not involved in this event. These data suggest that PI3K is required for the F‐actin‐based cytoskeleton changes during neutrophil rolling on E‐selectin, which may consequently regulate the rolling event. J. Cell. Biochem. 110: 910–919, 2010. © 2010 Wiley‐Liss, Inc.     

The miR‐3127‐5p/p‐STAT3 axis up‐regulates PD‐L1 inducing chemoresistance in non‐small‐cell lung cancer

It is less known about miRNA3127‐5p induced up‐regulation of PD‐L1, immune escape and drug resistance caused by increased PD‐L1 in lung cancer. In this study, lentivirus was transduced into lung cancer cells, and quantitative PCR and Western blot were used to detect the expression of PD‐L1. Then immunofluorescence assay was applied to detect autophagy, finally we explored the relationship between PD‐L1 expressions and chemoresistance in patients. As a result, we found that microRNA‐3127‐5p promotes pSTAT3 to induce the expression of PD‐L1; microRNA‐3127‐5p promotes STAT3 phosphorylation through suppressing autophagy, and autophagy could retaine pSTAT3 into the nucleus in miRNA‐3127‐5p knocked cells, and immune escape induced by elevated level of PD‐L1 results in chemoresistance of lung cancer. In conclusion, microRNA‐3127‐5p induces PD‐L1 elevation through regulating pSTAT3 expression. We also demonstrate that immune escape induced by PD‐L1 can be dismissed by corresponding monoclonal antibody.     

Light induces jasmonate‐isoleucine conjugation via OsJAR1‐dependent and ‐independent pathways in rice

The bioactive form of jasmonate is the conjugate of the amino acid isoleucine (Ile) with jasmonic acid (JA), which is biosynthesized in a reaction catalysed by the GH3 enzyme JASMONATE RESISTANT 1 (JAR1). We examined the biochemical properties of OsJAR1 and its involvement in photomorphogenesis of rice (Oryza sativa). OsJAR1 has a similar substrate specificities as its orthologue in Arabidopsis. However, osjar1 loss‐of‐function mutants did not show as severe coleoptile phenotypes as the JA‐deficient mutants coleoptile photomorphogenesis 2 (cpm2) and hebiba, which develop long coleoptiles in all light qualities we examined. Analysis of hormonal contents in the young seedling stage revealed that osjar1 mutants are still able to synthesize JA‐Ile conjugate in response to blue light, suggesting that a redundantly active enzyme can conjugate JA and Ile in rice seedlings. A good candidate for this enzyme is OsJAR2, which was found to be able to catalyse the conjugation of JA with Ile as well as with some additional amino acids. In contrast, if plants in the vegetative stage were mechanically wounded, the content of JA‐Ile was severely reduced in osjar1, demonstrating that OsJAR1 is the most important JA‐Ile conjugating enzyme in the wounding response during the vegetative stage.     

Seizure‐induced impairment in neuronal ketogenesis: Role of zinc‐α2‐glycoprotein in mitochondria

Ketone bodies (KBs) were known to suppress seizure. Untraditionally, neurons were recently reported to utilize fatty acids and produce KBs, but the effect of seizure on neuronal ketogenesis has not been researched. Zinc‐α2‐glycoprotein (ZAG) was reported to suppress seizure via unclear mechanism. Interestingly, ZAG was involved in fatty acid β‐oxidation and thus may exert anti‐epileptic effect by promoting ketogenesis. However, this promotive effect of ZAG on neuronal ketogenesis has not been clarified. In this study, we performed immunoprecipitation and mass spectrometry to identify potential interaction partners with ZAG. The mechanisms of how ZAG translocated into mitochondria were determined by quantitative coimmunoprecipitation after treatment with apoptozole, a heat shock cognate protein 70 (HSC70) inhibitor. ZAG level was modulated by lentivirus in neurons or adeno‐associated virus in rat brains. Seizure models were induced by magnesium (Mg²⁺)‐free artificial cerebrospinal fluid in neurons or intraperitoneal injection of pentylenetetrazole kindling in rats. Ketogenesis was determined by cyclic thio‐NADH method in supernatant of neurons or brain homogenate. The effect of peroxisome proliferator–activated receptor γ (PPARγ) on ZAG expression was examined by Western blot, quantitative real‐time polymerase chain reaction (qRT‐PCR) and chromatin immunoprecipitation qRT‐PCR. We found that seizure induced ketogenesis deficiency via a ZAG‐dependent mechanism. ZAG entered mitochondria through a HSC70‐dependent mechanism, promoted ketogenesis by binding to four β‐subunits of long‐chain L‐3‐hydroxyacyl‐CoA dehydrogenase (HADHB) and alleviated ketogenesis impairment in a neuronal seizure model and pentylenetetrazole‐kindled epileptic rats. Additionally, PPARγ activation up‐regulated ZAG expression by binding to promoter region of AZGP1 gene and promoted ketogenesis through a ZAG‐dependent mechanism.