A specific isozyme of 2'-5' oligoadenylate synthetase is a dual function proapoptotic protein of the Bcl-2 family

2-5(A) synthetases are a family of interferon-induced enzymes that polymerize ATP into 2'-5' linked oligoadenylates that activate RNase L and cause mRNA degradation. Because they all can synthesize 2-5(A), the reason for the existence of so many synthetase isozymes is unclear. Here we report that the 9-2 isozyme of 2-5(A) synthetase has an additional activity: it promotes apoptosis in mammalian cells. The proapoptotic activity of 9-2 was isozyme-specific and enzyme activity-independent. The 9-2-expressing cells exhibited many properties of cells undergoing apoptosis, such as DNA fragmentation, caspase activation, and poly ADP-ribose polymerase and lamin B cleavage. The isozyme-specific carboxyl-terminal tail of the 9-2 protein was shown, by molecular modeling, to contain a Bcl-2 homology 3 (BH3) domain, suggesting that it may be able to interact with members of the Bcl-2 family that contain BH1 and BH2 domains. Co-immunoprecipitate assays and confocal microscopy showed that 9-2 can indeed interact with the anti-apoptotic proteins Bcl-2 and Bclx(L) in vivo and in vitro. Mutations in the BH3 domain that eliminated the 9-2-Bcl-2 amd 9-2-Bclx(L) interactions also eliminated the apoptotic activity of 9-2. Thus, we have identified an interferon-induced dual function protein of the Bcl-2 family that can synthesize 2-5(A) and promote cellular apoptosis independently. Moreover, the cellular abundance of this protein is regulated by alternative splicing; the other isozymes encoded by the same gene are not proapoptotic.     

RecA realigns suboptimally paired frames of DNA repeats through a process that requires ATP hydrolysis

Microsatellite repeats such as mono-, di-, and trinucleotides are highly abundant and viable targets for homologous recombination in the genome. However, if recombination ensues in such repetitive regions, they are intrinsically prone to frame misalignments during pairing and might eventually give rise to genetic instabilities. Suboptimally paired frames lead to an abrogation of branch migration at the junctions of mixed sequences and repeats, due to a heterologous register. If so, can recombination machinery rectify such misalignments in order to avoid subsequent arrest in branch migration? We analyzed Escherichia coli RecA, the universal prototype of a recombinase, for its pairing abilities across repeats. We used a complementary pairing assay to test whether RecA can mediate realignments of stochastically paired suboptimal frames to a maximally aligned register. Here, we demonstrate that RecA-single stranded DNA filament indeed facilitates such a realignment, probably by sliding the paired strands across mono- and di- as well as trinucleotide repeats. These realignments apparently have no net directional bias. Such a putative "motor" function of RecA seems to be ATP hydrolysis-dependent.     

Circular RNA UBE2Q2 promotes malignant progression of gastric cancer by regulating signal transducer and activator of transcription 3-mediated autophagy and glycolysis

Gastric cancer remains the third leading cause of cancer-related mortality worldwide. Emerging evidence has shown that circular RNAs (circRNAs) play a critical regulatory role in the occurrence and development of various cancers through sponging miRNAs or acting as RNA-binding protein (RBP) sponges. We found that circUBE2Q2 was significantly upregulated in GC tissues and cell lines. Knockdown of circUBE2Q2 inhibited proliferation, migration, invasion, and glycolysis, and increased autophagy in vitro. In addition, knockdown of circUBE2Q2 inhibited GC tumorigenicity and metastasis potential in vivo. A series of experiments were performed to confirm that circUBE2Q2 regulates GC progression via the circUBE2Q2-miR-370-3p-STAT3 axis and promotes tumor metastasis through exosomal communication. Further in vivo experiments confirmed that, combination treatment of circUBE2Q2 knocking down and STAT3 inhibitor has synergistic effects on the gastric cancer growth inhibition, which provides a possibility to enhance the sensitivity of targeted drugs to gastric cancer through targeting circUBE2Q2. Our findings revealed that circUBE2Q2 may serve as a new proliferation-promoting factor and prognostic marker in gastric cancer.Subject terms: Gastric cancer, Gastric cancer     

Particle-bubble attachment in yeast flotation by colloidal gas aphrons

Flotation of microorganisms by colloidal gas aphrons (CGAs) is recognised as an inexpensive and effective method of separation. CGAs are micro-sized gas bubbles of 25 7m in average diameter and each is encapsulated in an aqueous shell of surfactant solution. High flotation efficiency can be achieved when colloidal gas aphrons are employed in a dilute suspension containing yeast cells. Under certain experimental conditions, the adsorption of yeast on the aphrons follows the Langmuir model. With changes in the pH and feed concentration, the mechanism of bubble attachment and detachment changes from a monolayer to a multilayer adsorption.     

Chromosome homology in the climbing rats,genus tylomys (Rodentia: Cricetidae)

The climbing rats (Tylomys spp.) have diploid numbers of 52 (T. panamensis), 42 (T. nudicaudus), 40 (T. n. gymnurus) and 36 (T. n. villai). Using G-band analysis we found that the variations are mainly of the Bobertsonian type, and practically all changes can be traced. G-banding also revealed that biarmed chromosomes with similar morphology may be composed of different components. Such conclusions were verified also by analysis of the chromosomes of interspecific hybrids. C-band staining showed that the constitutive heterochromatin of Tylomys is mainly located in the centromeric regions and the sex chromosomes, a situation similar to that of Microtus agrestis. In one specimen of T. panamensis, however, an additional terminal heterochromatic segment was found in one member of the large metacentric pair. Our data underline that in mammalian cytotaxonomy studies both C- and G-band (or C- and Q-band) techniques must be applied to gain maximal information.     

癌基因iASPP的克隆、表达与纯化

iASPP是新近发现的高度保守的p53相关基因,其蛋白产物定位于细胞核内,具有结合NFκBp65亚基和p53功能,进而抑制NFκB的转录调节和p53对凋亡的调节功能。利用RTPCR方法从人白血病细胞系U937中克隆出癌基因iASPP,将其克隆至原核表达载体pET28a(+)中,成功构建iASPP表达载体PIAF,重组质粒读码框和序列与预期一致。在IPTG诱导下,重组载体大肠杆菌Rosetta(DE3)菌株,表达产物经SDSPAGE和Westernblot方法分析证实系iASPP融合蛋白。利用Ni离子鳌合层析的方法纯化iASPP融合蛋白,经SDSPAGE鉴定其纯度超过80%。     

2��-O Methylation of the Viral mRNA Cap by West Nile Virus Evades Ifit1-Dependent and -Independent Mechanisms of Host Restriction In Vivo

Prior studies have shown that 2′-O methyltransferase activity of flaviviruses, coronaviruses, and poxviruses promotes viral evasion of Ifit1, an interferon-stimulated innate immune effector protein. Viruses lacking 2′-O methyltransferase activity exhibited attenuation in primary macrophages that was rescued in cells lacking Ifit1 gene expression. Here, we examined the role of Ifit1 in restricting pathogenesis in vivo of wild type WNV (WNV-WT) and a mutant in the NS5 gene (WNV-E218A) lacking 2′-O methylation of the 5′ viral RNA cap. While deletion of Ifit1 had marginal effects on WNV-WT pathogenesis, WNV-E218A showed increased replication in peripheral tissues of Ifit1 ^−/− mice after subcutaneous infection, yet this failed to correlate with enhanced infection in the brain or lethality. In comparison, WNV-E218A was virulent after intracranial infection as judged by increased infection in different regions of the central nervous system (CNS) and a greater than 16,000-fold decrease in LD₅₀ values in Ifit1 ^−/− compared to wild type mice. Ex vivo infection experiments revealed cell-type specific differences in the ability of an Ifit1 deficiency to complement the replication defect of WNV-E218A. In particular, WNV-E218A infection was impaired in both wild type and Ifit1 ^−/− brain microvascular endothelial cells, which are believed to participate in blood-brain barrier (BBB) regulation of virus entry into the CNS. A deficiency of Ifit1 also was associated with increased neuronal death in vivo, which was both cell-intrinsic and mediated by immunopathogenic CD8⁺ T cells. Our results suggest that virulent strains of WNV have largely evaded the antiviral effects of Ifit1, and viral mutants lacking 2′-O methylation are controlled in vivo by Ifit1-dependent and -independent mechanisms in different cell types.     

Survival adaptations of threeEuphorbia spp. in arid ecosystem

Three species ofEuphorbia (E. granulata, E. prostrata andE. hirta) exhibited both morphological and physiological adaptations in dry environmental conditions. High bound water, thick cuticle, dense hair covering and low stomatal index are some of the survival adaptations they developed under water stress conditions of the Indian arid zone.