XBP-1u suppresses autophagy by promoting the degradation of FoxO1 in cancer cells

Autophagy is activated to maintain cellular energy homeostasis in response to nutrient starvation. However, autophagy is not persistently activated, which is poorly understood at a mechanistic level. Here, we report that turnover of FoxO1 is involved in the dynamic autophagic process caused by glutamine starvation. X-box-binding protein-1u (XBP-1u) has a critical role in FoxO1 degradation by recruiting FoxO1 to the 20S proteasome. In addition, the phosphorylation of XBP-1u by extracellular regulated protein kinases1/2 (ERK1/2) on Ser61 and Ser176 was found to be critical for the increased interaction between XBP-1u and FoxO1 upon glutamine starvation. Furthermore, knockdown of XBP-1u caused the sustained level of FoxO1 and the persistent activation of autophagy, leading to a significant decrease in cell viability. Finally, the inverse correlation between XBP-1u and FoxO1 expression agrees well with the expression profiles observed in many human cancer tissues. Thus, our findings link the dynamic process of autophagy to XBP-1u-induced FoxO1 degradation.     

Design,synthesis and biological evaluation of novel 1H-1,2,4-triazole,benzothiazole and indazole-based derivatives as potent FGFR1 inhibitors viafragment-based virtual screening

Abstract

Fibroblast growth-factor receptor (FGFR) is a potential target for cancer therapy. We designed three novel series of FGFR1 inhibitors bearing indazole, benzothiazole, and 1H-1,2,4-triazole scaffold via fragment-based virtual screening. All the newly synthesised compounds were evaluated in vitro for their inhibitory activities against FGFR1. Compound 9d bearing an indazole scaffold was first identified as a hit compound, with excellent kinase inhibitory activity (IC₅₀ = 15.0?nM) and modest anti-proliferative activity (IC₅₀ = 785.8?nM). Through two rounds of optimisation, the indazole derivative 9?u stood out as the most potent FGFR1 inhibitors with the best enzyme inhibitory activity (IC₅₀ = 3.3?nM) and cellular activity (IC₅₀ = 468.2?nM). Moreover, 9?u also exhibited good kinase selectivity. In addition, molecular docking study was performed to investigate the binding mode between target compounds and FGFR1.     

An in vitro network of intermolecular interactions between viral RNA segments of an avian H5N2 influenza A virus: comparison with a human H3N2 virus

The genome of influenza A viruses (IAV) is split into eight viral RNAs (vRNAs) that are encapsidated as viral ribonucleoproteins. The existence of a segment-specific packaging mechanism is well established, but the molecular basis of this mechanism remains to be deciphered. Selective packaging could be mediated by direct interaction between the vRNA packaging regions, but such interactions have never been demonstrated in virions. Recently, we showed that the eight vRNAs of a human H3N2 IAV form a single interaction network in vitro that involves regions of the vRNAs known to contain packaging signals in the case of H1N1 IAV strains. Here, we show that the eight vRNAs of an avian H5N2 IAV also form a single network of interactions in vitro, but, interestingly, the interactions and the regions of the vRNAs they involve differ from those described for the human H3N2 virus. We identified the vRNA sequences involved in five of these interactions at the nucleotide level, and in two cases, we validated the existence of the interaction using compensatory mutations in the interacting sequences. Electron tomography also revealed significant differences in the interactions taking place between viral ribonucleoproteins in H5N2 and H3N2 virions, despite their canonical ‘7 + 1’ arrangement.     

Mutations in IMPG2, Encoding Interphotoreceptor Matrix Proteoglycan 2, Cause Autosomal-Recessive Retinitis Pigmentosa

Retinitis pigmentosa (RP) is a heterogeneous group of inherited retinal diseases caused by progressive degeneration of the photoreceptor cells. Using autozygosity mapping, we identified two families, each with three affected siblings sharing large overlapping homozygous regions that harbored the IMPG2 gene on chromosome 3. Sequence analysis of IMPG2 in the two index cases revealed homozygous mutations cosegregating with the disease in the respective families: three affected siblings of Iraqi Jewish ancestry displayed a nonsense mutation, and a Dutch family displayed a 1.8 kb genomic deletion that removes exon 9 and results in the absence of seven amino acids in a conserved SEA domain of the IMPG2 protein. Transient transfection of COS-1 cells showed that a construct expressing the wild-type SEA domain is properly targeted to the plasma membrane, whereas the mutant lacking the seven amino acids appears to be retained in the endoplasmic reticulum. Mutation analysis in ten additional index cases that were of Dutch, Israeli, Italian, and Pakistani origin and had homozygous regions encompassing IMPG2 revealed five additional mutations; four nonsense mutations and one missense mutation affecting a highly conserved phenylalanine residue. Most patients with IMPG2 mutations showed an early-onset form of RP with progressive visual-field loss and deterioration of visual acuity. The patient with the missense mutation, however, was diagnosed with maculopathy. The IMPG2 gene encodes the interphotoreceptor matrix proteoglycan IMPG2, which is a constituent of the interphotoreceptor matrix. Our data therefore show that mutations in a structural component of the interphotoreceptor matrix can cause arRP.     

DNA barcoding analysis of Coleoidea (Mollusca: Cephalopoda) from Chinese waters

Coleoids are part of the Cephalopoda class, which occupy an important position in most oceans both at an ecological level and at a commercial level. Nevertheless, some coleoid species are difficult to distinguish with traditional morphological identification in cases when specimens are heavily damaged during collection or when closely related taxa are existent. As a useful tool for rapid species assignment, DNA barcoding may offer significant potential for coleoid identification. Here, we used two mitochondrial fragments, cytochrome c oxidase I and the large ribosomal subunit (16S rRNA), to assess whether 34 coleoids accounting for about one-third of the Chinese coleoid fauna could be identified by DNA barcoding technique. The pairwise intra- and interspecific distances were assessed, and relationships among species were estimated by NJ and bayesian analyses. High levels of genetic differentiation within Loliolus beka led to an overlap between intra- and interspecific distances. All remaining species forming well-differentiated clades in the NJ and bayesian trees were identical for both fragments. Loliolus beka possessed two mitochondrial lineages with high levels of intraspecific distances, suggesting the occurrence of cryptic species. This study confirms the efficacy of DNA barcoding for identifying species as well as discovering cryptic diversity of Chinese coleoids. It also lays a foundation for other ecological and biological studies of Coleoidea.     

Cell density-dependent reduction of dihydroceramide desaturase activity in neuroblastoma cells

We applied a metabolic approach to investigate the role of sphingolipids in cell density-induced growth arrest in neuroblastoma cells. Our data revealed that sphingolipid metabolism in neuroblastoma cells significantly differs depending on the cells' population context. At high cell density, cells exhibited G0/G1 cell-cycle arrest and reduced ceramide, monohexosylceramide, and sphingomyelin, whereas dihydroceramide was significantly increased. In addition, our metabolic-labeling experiments showed that neuroblastoma cells at high cell density preferentially synthesized very long chain (VLC) sphingolipids and dramatically decreased synthesis of sphingosine-1-phosphate (S1P). Moreover, densely populated neuroblastoma cells showed increased message levels of both anabolic and catabolic enzymes of the sphingolipid pathway. Notably, our metabolic-labeling experiments indicated reduced dihydroceramide desaturase activity at confluence, which was confirmed by direct measurement of dihydroceramide desaturase activity in situ and in vitro. Importantly, we could reduce dihydroceramide desaturase activity in low-density cells by applying conditional media from high-density cells, as well as by adding reducing agents, such as DTT and L-cysteine to the media. In conclusion, our data suggest a role of the sphingolipid pathway, dihydroceramides desaturase in particular, in confluence-induced growth arrest in neuroblastoma cells.     

R-Ras and Rac GTPase cross-talk regulates hematopoietic progenitor cell migration, homing, and mobilization

Adult hematopoietic progenitor cells (HPCs) are maintained by highly coordinated signals in the bone marrow. The molecular mechanisms linking intracellular signaling network of HPCs with their microenvironment remain poorly defined. The Rho family GTPase Rac1/Rac2 has previously been implicated in cell functions involved in HPC maintenance, including adhesion, migration, homing, and mobilization. In the present studies we have identified R-Ras, a member of the Ras family, as a key signal mediator required for Rac1/Rac2 activation. We found that whereas Rac1 activity is up-regulated upon stem cell factor, integrin, or CXCL12 stimulation, R-Ras activity is inversely up-regulated. Expression of a constitutively active R-Ras mutant resulted in down-regulation of Rac1-activity whereas deletion of R-Ras led to an increase in Rac1/Rac2 activity and signaling. R-Ras(-/-) HPCs displayed a constitutively assembled cortical actin structure and showed increased directional migration. Rac1/Rac2 inhibition reversed the migration phenotype of R-Ras(-/-) HPCs, similar to that by expressing an R-Ras active mutant. Furthermore, R-Ras(-/-) mice showed enhanced responsiveness to G-CSF for HPC mobilization and exhibited decreased bone marrow homing. Transplantation experiments indicate that the R-Ras deficiency-induced HPC mobilization is a HPC intrinsic property. These results indicate that R-Ras is a critical regulator of Rac signaling required for HPC migration, homing, and mobilization.     

SAR studies of 1,5-diarylpyrazole-based CCK1 receptor antagonists

A high throughput screening campaign revealed compound 1 as a potent antagonist of the human CCK(1) receptor. Here, we report the syntheses and SAR studies of 1,5-diarylpyrazole analogs with various structural modifications of the alkane side chain of the molecule. The difference in affinity between the two enantiomers for the CCK(1) receptor and the flexible nature of the linker led to the design of constrained analogs with increased potency.