Structural basis for interaction between the Ubp3 deubiquitinating enzyme and its Bre5 cofactor

The Bre5 protein is a cofactor for the deubiquitinating enzyme Ubp3, and it contains a nuclear transfer factor 2 (NTF2)-like protein recognition module that is essential for Ubp3 activity. In this study, we report the x-ray crystal structure of the Bre5 NTF2-like domain and show that it forms a homodimeric structure that is similar to other NTF2-like domains, except for the presence of an intermolecular disulfide bond in the crystals. Sedimentation equilibrium studies reveal that under non-reducing conditions, the Bre5 NTF2-like domain is exclusively dimeric, whereas a disulfide bond-deficient mutant undergoes a monomer-dimer equilibrium with a dissociation constant in the midnanomolar range, suggesting that dimer formation and possibly also disulfide bond formation may modulate Bre5 function in vivo. Using deletion analysis, we also identify a novel N-terminal domain of Ubp3 that is necessary and sufficient for interaction with Bre5 and use isothermal titration calorimetry to show that Bre5 and Ubp3 form a 2:1 complex, in contrast to other reported NTF2-like domain/protein interactions that form 1:1 complexes. Finally, we employ structure-based mutagenesis to map the Ubp3 binding surface of Bre5 to a region near the Bre5 dimer interface and show that this binding surface of Bre5 is important for Ubp3 function in vivo. Together, these studies provide novel insights into protein recognition by NTF2-like domains and provide a molecular scaffold for understanding how Ubp3 function is regulated by Bre5 cofactor binding.     

Characterization and phytochemical constituents of Periploca hydaspidis Falc crude extract and its anticancer activities

The current study aims to investigate the anticancer potential of Periploca hydaspidis extracts against HCCLM3 and MDA-MB 231 cell lines with invasive properties and to identify molecular targets underlying its action mechanism. Cytotoxic screening of plant extracts was done via MTT assay against liver and breast cancer cell lines and GC/MS of the best cytotoxic fraction was performed to identify its chemical composition. Flow cytometry detected apoptosis and cell-cycle changes after drug treatment. The specified cells were studied for migration and invasion potential along with performing western blot analysis of proteins involved in apoptosis, cell-cycle, metastasis, and MAPK (Mitogen-activated protein kinase) cell-signaling pathway. The results revealed the crude methanol (PHM) fraction of P. hydaspidis shown dose and time dependent cell-proliferative inhibition response. GC/MS analysis detected 54 compounds of which fatty acids (29.8%), benzenoids (15.7%), and esters (14.3%) constituted the bulk. The inhibitory effect against cancer cells was linked with cell-cycle arrest at G0/G1 phase, induction of apoptosis, reduced migration and invasion capabilities post treatment. PHM induced apoptosis via downregulation of anti-apoptotic (survivin, B-cell lymphoma Extra-large; BCL-XL, X-linked inhibitor of apoptosis protein; XIAP, Myelocytomatosis; C-myc), metastatic (Matrix metallopeptidases 9/2; MMP9/2), and cell-cycle regulatory (cyclin D1 and E) proteins, whereas upregulation of pro-apoptotic proteins (Bcl-2 homologous antagonist/killer; BAK, Bcl-2-Associate X protein; BAX, cleaved caspases; 3,7,8,9, and PARP) and activation of MAPK (Jun amino-terminal kinase; JNK and P38) pathway. P38 was needed for PHM-induced apoptosis, where the inhibition of P38 by pharmacological inhibitor (SB239063) diminished the apoptotic effects. Overall, our results conclude that PHM can inhibit cell-proliferation and induce apoptotic effects by activation of P38 MAPK cell-signaling pathway. This suggests the methanol fraction of P. hydaspidis (PHM) to have anticancer compounds, potentially useful for treating liver and breast cancer. In future, one-step advance studies of PHM regarding its role in metastatic inhibition, immune response modulation for reducing tumor, and inducing apoptosis in suitable animal models would be an interesting and promising research area.     

Molecular characterization and expression of cyclic nucleotide gated ion channels 19 and 20 in Arabidopsis thaliana for their potential role in salt stress

Cyclic nucleotide gated ion channels (CNGCs) in plants have very important role in signaling and development. The study reports role of CNGC19 and CNGC20 in salt stress in A. thaliana. In-silico, genome wide analysis showed that CNGC19 and CNGC20 are related to salt stress with maximum expression after 6 h in A. thaliana. The position of inserted T-DNA was determined (in-vivo) through TAIL-PCR for activation tagged mutants of CNGC19 and CNGC20 under salt stress. The expression of AtCNGC19 and AtCNGC20 after cloning under 35S promoter of expression vectors pBCH1 and pEarleyGate100 was determined in A. thaliana by real-time PCR analysis. Genome wide analysis showed that AtCNGC11 had lowest and AtCNGC20 highest molecular weight as well as number of amino acid residues. In-vivo expression of AtCNGC19 and AtCNGC20 was enhanced through T-DNA insertion and 35S promoter in over-expressed plants under high salt concentration. AtCNGC19 was activated twice in control and about five times under 150 mM NaCl stress level, and expression value was also higher than AtCNGC20. Phenotypically, over-expressed plants and calli were healthier while knock-out plants and calli showed retarded growth under salinity stress. The study provides new insight for the role of AtCNGC19 and AtCNGC20 under salt stress regulation in A. thaliana and will be helpful for improvement of crop plants for salt stress to combat food shortage and security.     

Carbon Quantum Dots as Multi-Purpose Nanomaterial in Stem Cell Therapy

During stem cell therapy, some issues, such as obscure fate of stem cells or their low survival rate in the body, should be addressed to boost their therapeutic efficiency. Nanotechnology offers a suitable solution to combat such limitations. Carbon quantum dots (CQDs) are carbon-based nanomaterials and may be used as multi-purpose compounds in stem cell therapy. CQDs are excellent choices for stem cell labeling thanks to their special features such as optical properties and good biocompatibility. Besides, they can modulate the biological function of stem cells, such as their proliferation, homing ability, and differentiation properties. Considering the charismatic feature of CQDs and their broad unique effect on stem cells, the current review aims to summarize the most advancements in this field. Hence, we first focused on CQDs synthesis and their applications. In the next section, the stem cell categories will be discussed, and the final part is dedicated to the recent research evaluating the impact of CQDs on stem cell therapy.     

Effect of Red Bull energy drink on cardiovascular and renal function

Energy drink consumption has been anecdotally linked to the development of adverse cardiovascular effects in consumers, although clinical trials to support this link are lacking. The effects of Red Bull^® energy drink on cardiovascular and neurologic functions were examined in college-aged students enrolled at Winona State University. In a double-blind experiment where normal calorie and low calorie Red Bull^® were compared to normal and low calorie placebos, no changes in overall cardiovascular function nor blood glucose (mg/dL) were recorded in any participant (n = 68) throughout a 2-h test period. However, in the second experiment, nine male and twelve female participants subjected to a cold pressor test (CPT) before and after Red Bull^® consumption showed a significant increase in blood sugar levels pre- and post Red Bull^® consumption. There was a significant increase in diastolic blood pressure of the male volunteers immediately after submersion of the hand in the 5°C water for the CPT. Under the influence of Red Bull^®, the increase in diastolic pressure for the male participants during the CPT was negated. There were no significant changes in the blood pressure of the female participants for the CPT with or without Red Bull^®. Finally, the CPT was used to evaluate pain threshold and pain tolerance before and after Red Bull^® consumption. Red Bull^® consumption was associated with a significant increase in pain tolerance in all participants. These findings suggest that Red Bull^® consumption ameliorates changes in blood pressure during stressful experiences and increases the participants’ pain tolerance.     

Chromium tolerance,oxidative stress response,morphological characteristics,and FTIR studies of phytopathogenic fungus <Emphasis Type="Italic">Sclerotium rolfsii</Emphasis>

Sclerotium rolfsii is one of the most destructive fungal plant pathogens that can infect over 500 plants and can adapt to diverse environmental conditions. The present research work was carried out to evaluate the impact of both hexa- and trivalent chromium (Cr) on growth, morphology, enzymatic characteristics, and metal accumulation in S. rolfsii under laboratory conditions. Experiments were performed in both malt extract broth and agar growth medium amended with six different concentrations (10, 20, 40, 60, 80, and 100 ppm) of each Cr(III) and Cr(VI) ions inoculated with fungus and incubated for 6–7 days at 25 ± 3 °C. In broth medium, the total protein content was declined and activities of antioxidant enzymes were increased with an increase in metal concentrations. Lower concentrations (10 ppm) of the metal ions stimulated the growth of fungus and higher concentrations (60–100) inhibited it. The Fourier transform infrared spectroscopy (FTIR) assessment showed hydroxyl, carboxyl, and amine groups as major metal binding sites. In agar medium, tolerance index was decreased up to 0.56 at 10–80 ppm of Cr(III) and up to 0.62 at 10–60 ppm of Cr(VI). Considerable modifications were observed in hyphal and sclerotial morphology with an increase in concentration of metal ions. The current study concluded that interference of Cr with growth and physiological process of S. rolfsii could affect its infection level on its host plant. This study provides important information regarding cultivation of susceptible plant varieties in Cr-polluted soil as evidenced by pathogen growth up to 50 ppm of Cr(III) and Cr(VI) ions.     

Study of rat hypothalamic proteome by HPLC/ESI ion trap and HPLC/ESI‐Q‐TOF MS

The proteomic profile of hypothalamus, a key organ of CNS, is explored here by employing two widely used MS techniques, i.e. HPLC/ESI‐ion trap and HPLC/ESI‐quadrupole‐TOF MS. Strong cation exchange is used for the fractionation of peptides and protein search engine MASCOT is employed for data query. One hundred and thirty six proteins with 10 973 peptides were identified by HPLC/ESI‐ion trap MS, while 140 proteins with 32 183 peptides were characterized by HPLC/ESI‐quadrupole‐TOF MS. Among the total 198 proteins identified in both experiments, 78 proteins were common in both sets of conditions. The rest of the 120 proteins were identified distinctly in both MS strategies, i.e. 58 unique proteins were found using the quadrupole‐TOF while 62 were found with the HPLC/ESI‐ion trap. Moreover, these proteins were classified into groups based on their functions performed in the body. Results presented here identified some important signal and cellular defense proteins inevitable for survival in stressed conditions. Additionally, it is also shown that any single MS strategy is not reliable for good results due to loss of data depending on sensitivity of the instrument used.     

Delayed expansion and contraction of CD8+ T cell response during infection with virulent Salmonella typhimurium

Ag presentation to CD8(+) T cells often commences immediately after infection, which facilitates their rapid expansion and control of infection. Subsequently, the primed cells undergo rapid contraction. We report that this paradigm is not followed during infection with virulent Salmonella enterica, serovar Typhimurium (ST), an intracellular bacterium that replicates within phagosomes of infected cells. Although susceptible mice die rapidly (approximately 7 days), resistant mice (129 x 1SvJ) harbor a chronic infection lasting approximately 60-90 days. Using rOVA-expressing ST (ST-OVA), we show that T cell priming is considerably delayed in the resistant mice. CD8(+) T cells that are induced during ST-OVA infection undergo delayed expansion, which peaks around day 21, and is followed by protracted contraction. Initially, ST-OVA induces a small population of cycling central phenotype (CD62L(high)IL-7Ralpha(high)CD44(high)) CD8(+) T cells. However, by day 14-21, majority of the primed CD8(+) T cells display an effector phenotype (CD62L(low)IL-7Ralpha(low)CD44(high)). Subsequently, a progressive increase in the numbers of effector memory phenotype cells (CD62L(low)IL-7Ralpha(high)CD44(high)) occurs. This differentiation program remained unchanged after accelerated removal of the pathogen with antibiotics, as majority of the primed cells displayed an effector memory phenotype even at 6 mo postinfection. Despite the chronic infection, CD8(+) T cells induced by ST-OVA were functional as they exhibited killing ability and cytokine production. Importantly, even memory CD8(+) T cells failed to undergo rapid expansion in response to ST-OVA infection, suggesting a delay in T cell priming during infection with virulent ST-OVA. Thus, phagosomal lifestyle may allow escape from host CD8(+) T cell recognition, conferring a survival advantage to the pathogen.     

Study of interaction effect between triacontanol and nitric oxide on alleviating of oxidative stress arsenic toxicity in coriander seedlings

In this study, triacontanol (TRIA) and nitric oxide (NO) interaction on arsenic (As)-induced oxidative stress tolerance in coriander (Coriandrum sativum L.) plants was investigated. The results showed that As had a significant adverse effect on the plant’s biomass. The seedlings pretreated with TRIA and NO significantly increased growth reduction induced by the metalloid. The obtained results indicated that the application of TRIA and sodium nitroprusside (SNP) generally reduced oxidative markers such as of electrolyte leakage percentage, malondialdehyde and H₂O₂ contents under As toxicity, while application of As treatment without TRIA?+?SNP increased these oxidative parameters compared to the control. The non-enzymatic antioxidant contents such as total phenol, anthocyanin, carotenoid, ascorbic acid and reduced glutathione (GSH) were extracted and assayed from both control and treated plants. It was found that TRIA?+?SNP treatments have a profound effect on the antioxidant metabolism and caused an enhancement in non-enzymatic antioxidant potentials under As toxicity in coriander. Moreover, the results revealed a mutually amplifying reaction between TRIA and NO in reducing As-induced damages.