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Abstract : Autophagy is a lysosome-dependent intracellular degradation process that allows recycling of cytoplasmic constituents into bioenergetic and biosynthetic materials for maintenance of homeostasis. Since the function of autophagy is particularly important in various stress conditions, perturbation of autophagy can lead to cellular dysfunction and diseases. Accumulation of abnormal protein aggregates, a common cause of neurodegenerative diseases, can be reduced through autophagic degradation. Recent studies have revealed defects in autophagy in most cases of neurodegenerative disorders. Moreover, deregulated excessive autophagy can also cause neurodegeneration. Thus, healthy activation of autophagy is essential for therapeutic approaches in neurodegenerative diseases and many autophagy-regulating compounds are under development for therapeutic purposes. This review describes the overall role of autophagy in neurodegeneration, focusing on various therapeutic strategies for modulating specific stages of autophagy and on the current status of drug development.

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Abstract : Degradation of chlorophyll (Chl) by Chl catabolic enzymes (CCEs) causes the loss of green color that typically occurs during senescence of leaves. In addition to CCEs, STAYGREEN1 (SGR1) functions as a key regulator of Chl degradation. Although sgr1 mutants in many plant species exhibit a stay-green phenotype, the biochemical function of the SGR1 protein remains elusive. Many recent studies have examined the physiological and molecular roles of SGR1 and its homologs (SGR2 and SGR-LIKE) in Chl metabolism, finding that these proteins have different roles in different species. In this review, we summarize the recent studies on SGR and discuss the most likely functions of SGR homologs.

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Abstract : Nuclear factor erythroid 2-related factor 2 (Nrf2) is an important redox-sensitive transcription factor that regulates the expression of several cytoprotective genes. More recently, genetic analyses of human tumors have indicated that Nrf2 may cause resistance to chemotherapy. In this study, we found that the expression levels of Nrf2 and its target genes GCLC, HO-1, NQO1 were significantly higher in cisplatin-resistant A549 (A549/CDDP) cells than those in A549 cells, and this resistance was partially reversed by Nrf2 siRNA. 3′,4′,5′,5,7-Pentamethoxyflavone (PMF), a natural flavonoid extracted from Rutaceae plants, sensitized A549/CDDP to CDDP and substantially induced apoptosis compared with that of CDDP alone treated group, and this reversal effect decreased when Nrf2 was downregulated by siRNA. Mechanistically, PMF reduced Nrf2 expression leading to a reduction of Nrf2 downstream genes, and in contrast, this effect was decreased by blocking Nrf2 with siRNA. Taken together, these results demonstrated that PMF could be used as an effective adjuvant sensitizer to increase the efficacy of chemotherapeutic drugs by downregulating Nrf2 signaling pathway.

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Abstract : Protein O-GlcNAcylation, dictated by cellular UDP-N-acetylglucosamine (UDP-GlcNAc) levels, plays a crucial role in posttranslational modifications. The enzyme GlcNAc kinase (NAGK, E.C. 2.7.1.59) catalyzes the formation of GlcNAc-6-phosphate, which is a major substrate for the biosynthesis of UDP-GlcNAc. Recent studies have revealed the expression of NAGK in different types of cells especially in neuronal dendrites. Here, by immunocytochemistry (ICC) and immunonucleochemistry (INC) of cultured rat hippocampal neurons, HEK293T and GT1-7 cells, we have showed that NAGK immuno-reactive punctae being present in the nucleoplasm colocalized with small nuclear ribonucleoprotein-associated protein N (snRNPN) and p54NRB, which are speckle and paraspeckle markers, respectively. Furthermore, NAGK IR cluster was also found to be colocalized with GTF2H5 (general transcription factor IIH, polypeptide 5) immuno reactive punctae. In addition, relative localization to the ring of nuclear lamin matrix and to GlcNAc, which is highly enriched in nuclear pore complexes, showed that NAGK surrounds the nucleus at the cytoplasmic face of the nuclear outer membrane. By in situ proximity ligation assay (PLA) we confirmed the colocalization of NAGK with snRNPN in the nucleus and in dendrites, while we also verified the interactions of NAGK with p54NRB, and with GTF2H5 in the nucleus. These associations between NAGK with speckle, paraspeckle and general transcription factor suggest its regulatory roles in gene expression.

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Abstract : Low-barrier hydrogen bonds (LBHBs) have been proposed to have important influences on the enormous reaction rate increases achieved by many enzymes. Δ⁵-3-ketosteroid isomerase (KSI) catalyzes the allylic isomerization of Δ⁵-3-ketosteroid to its conjugated Δ⁴-isomers at a rate that approaches the diffusion limit. Tyr14, a catalytic residue of KSI, has been hypothesized to form an LBHB with the oxyanion of a dienolate steroid intermediate generated during the catalysis. The unusual chemical shift of a proton at 16.8 ppm in the nuclear magnetic resonance spectrum has been attributed to an LBHB between Tyr14 Oη and C3-O of equilenin, an intermediate analogue, in the active site of D38N KSI. This shift in the spectrum was not observed in Y30F/Y55F/D38N and Y30F/Y55F/Y115F/D38N mutant KSIs when each mutant was complexed with equilenin, suggesting that Tyr14 could not form LBHB with the intermediate analogue in these mutant KSIs. The crystal structure of Y30F/Y55F/Y115F/D38N-equilenin complex revealed that the distance between Tyr14 Oη and C3-O of the bound steroid was within a direct hydrogen bond. The conversion of LBHB to an ordinary hydrogen bond in the mutant KSI reduced the binding affinity for the steroid inhibitors by a factor of 8.1?11. In addition, the absence of LBHB reduced the catalytic activity by only a factor of 1.7?2. These results suggest that the amount of stabilization energy of the reaction intermediate provided by LBHB is small compared with that provided by an ordinary hydrogen bond in KSI.

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Abstract : NF-E2-related factor 2 (Nrf2), a basic leucine zipper transcription factor, has recently received a great deal of attention as an important molecule that enhances antioxidative defenses and induces resistance to chemotherapy or radiotherapy. In this study, we investigated the apoptosis-inducing and Nrf2-upregulating effects of quercetin on malignant mesothelioma (MM) MSTO-211H and H2452 cells. Quercetin treatment inhibited cell growth and led to upregulation of Nrf2 at both the mRNA and protein levels without altering the ubiquitination and extending the half-life of the Nrf2 protein. Following treatment with quercetin, analyses of the nuclear level of Nrf2, Nrf2 antioxidant response element-binding assay, Nrf2 promoter-luc assay, and RT-PCR toward the Nrf2-regulated gene, heme oxygenase-1, demonstrated that the induced Nrf2 is transcriptionally active. Knockdown of Nrf2 expression with siRNA enhanced cytotoxicity due to the induction of apoptosis, as evidenced by an increase in the level of proapoptotic Bax, a decrease in the level of antiapoptotic Bcl-2 with enhanced cleavage of caspase-3 and PARP proteins, the appearance of a sub-G₀/G₁ peak in the flow cytometric assay, and increased percentage of apoptotic propensities in the annexin V binding assay. Effective reversal of apoptosis was observed following pretreatment with the pan-caspase inhibitor Z-VAD. Moreover, Nrf2 knockdown exhibited increased sensitivity to the anticancer drug, cisplatin, presumably by potentiating the oxidative stress induced by cisplatin. Collectively, our data demonstrate the importance of Nrf2 in cytoprotection, survival, and drug resistance with implications for the potential significance of targeting Nrf2 as a promising strategy for overcoming resistance to chemotherapeutics in MM.

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Abstract : Odin has been implicated in the downstream signaling pathway of receptor tyrosine kinases, such as the epidermal growth factor and Eph receptors. However, the physiologically relevant function of Odin needs to be further determined. In this study, we used Odin heterozygous mice to analyze the Odin expression pattern; the targeted allele contained a β-geo gene trap vector inserted into the 14th intron of the Odin gene. Interestingly, we found that Odin was exclusively expressed in ependymal cells along the brain ventricles. In particular, Odin was highly expressed in the subcommissural organ, a small ependymal glandular tissue. However, we did not observe any morphological abnormalities in the brain ventricles or ependymal cells of Odin null-mutant mice. We also generated BAC transgenic mice that expressed the PTB-deleted Odin (dPTB) after a floxed GFP-STOP cassette was excised by tissue-specific Cre expression. Strikingly, Odin-dPTB expression played a causative role in the development of the hydrocephalic phenotype, primarily in the midbrain. In addition, Odin-dPTB expression disrupted proper development of the subcommissural organ and interfered with ependymal cell maturation in the cerebral aqueduct. Taken together, our findings strongly suggest that Odin plays a role in the differentiation of ependymal cells during early postnatal brain development.

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Abstract : Human osteosarcoma usually presented a high tendency to metastatic spread and caused poor outcomes, however, the underlying mechanism was still largely unknown. In the present study, using a series of in vitro experiments and an animal model, we investigated the roles of HOX antisense intergenic RNA (HOTAIR) during the proliferation and invasion of osteosarcoma. According with our results, HOTAIR was commonly overexpressed in osteosarcoma, which significantly correlated with advanced tumor stage, highly histological grade and poor prognosis. In vitro and in vivo experiments demonstrated that knockdown of HOTAIR could notably suppress cellular proliferation, inhibit invasion and decrease the secretion of MMP2 and MMP9 in osteosarcoma. Collectively, our results suggested that HOTAIR might be a potent therapeutic target for osteosarcoma.

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Abstract : Recognition of cytosolic DNA initiates a series of innate immune responses by inducing IFN-I production and subsequent triggering JAK1-STAT1 signaling which plays critical roles in the pathogenesis of infection, inflammation and autoimmune diseases through promoting B cell activation and antibody responses. The stimulator of interferon genes protein (STING) has been demonstrated to be a critical hub of type I IFN induction in cytosolic DNA-sensing pathways. However, it still remains unknown whether cytosolic DNA can directly activate the JAK1-STAT1 signaling or not. And the role of STING is also unclear in this response. In the present study, we found that dsDNA directly triggered the JAK1-STAT1 signaling by inducing phosphorylation of the Lyn kinase. Moreover, this response is not dependent on type I IFN receptors. Interestingly, STING could inhibit dsDNA-triggered activation of JAK1-STAT1 signaling by inducing SHP-1 and SHP-2 phosphorylation. In addition, compared with normal B cells, the expression of STING was significantly lower and the phosphorylation level of JAK1 was significantly higher in B cells from MRL/lpr lupus-prone mice, highlighting the close association between STING low-expression and JAK1-STAT1 signaling activation in B cells in autoimmune diseases. Our data provide a molecular insight into the novel role of STING in dsDNA-mediated inflammatory disorders.

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Abstract : Obesity is the fifth leading risk for death globally, and a significant challenge to global health. It is a common, complex, non-malignant disease and develops due to interactions between the genes and the environment. DNA methylation can act as a downstream effector of environmental signals; analysis of this process therefore holds substantial promise for identifying mechanisms through which genetic and environmental factors jointly contribute to disease risk. To assess the effects of excessive weight and obesity on gene-specific methylation levels of promoter regions, we determined the methylation status of four genes involved in inflammation and oxidative stress [interleukin 6 (IL6), tumor necrosis factor α (TNFα), mitochondrial transcription factor A (TFAM), and glucose transport 4 (GLUT4)] in blood cell-derived DNA from healthy women volunteers with a range of body mass indices (BMIs) by methylation-specific PCR. Interestingly, the samples from obese individuals (BMI ≥ 30 kg/m²) showed significantly increased hypermethylation for IL6 gene compared to normal weight (BMI < 23 kg/m²) and overweight samples (23 kg/m² ≤ BMI < 30 kg/m²) (P = 0.034 and P = 0.026). However, there was no statistically significant difference in promoter methylation of the other 3 genes between each group. These findings suggest that aberrant DNA methylation of IL6 gene promoter may play an important role in the etiology and pathogenesis of obesity and IL6 methylation could be used as molecular biomarker for obesity risk assessment. Further studies are required to elucidate the potential mechanisms underlying this relationship.