Membrane Targeting and GTPase Activity of Rab7 Are Required for Its Ubiquitination by RNF167

Rab7 is a GTPase that controls late endosome and lysosome trafficking. Recent studies have demonstrated that Rab7 is ubiquitinated, a post-translational modification mediated by an enzymatic cascade. To date, only one ubiquitin E3 ligase and one deubiquitinase have been identified in regulating Rab7 ubiquitination. Here, we report that RNF167, a transmembrane endolysosomal ubiquitin ligase, can ubiquitinate Rab7. Using immunoprecipitation and in vitro ubiquitination assays, we demonstrate that Rab7 is a direct substrate of RNF167. Subcellular fractionation indicates that RNF167 activity maintains Rab7′s membrane localization. Epifluorescence microscopy in HeLa cells shows that Rab7-positive vesicles are larger under conditions enabling Rab7 ubiquitination by RNF167. Characterization of its ubiquitination reveals that Rab7 must be in its GTP-bound active form for membrane anchoring and, thus, accessible for RNF167-mediated ubiquitin attachment. Cellular distribution analyses of lysosome marker Lamp1 show that vesicle positioning is independent of Rab7 and RNF167 expression and that Rab7 endosomal localization is not affected by RNF167 knockdown. However, both Rab7 and RNF167 depletion affect each other’s lysosomal localization. Finally, this study demonstrates that the RNF167-mediated ubiquitination of Rab7 GTPase is impaired by variants of Charcot–Marie–Tooth Type 2B disease. This study identified RNF167 as a new ubiquitin ligase for Rab7 while expanding our knowledge of the mechanisms underlying the ubiquitination of Rab7.     

Mitochondrial Dysfunction Plays Central Role in Nonalcoholic Fatty Liver Disease

Nonalcoholic fatty liver disease (NAFLD) is a global pandemic that affects one-quarter of the world’s population. NAFLD includes a spectrum of progressive liver disease from steatosis to nonalcoholic steatohepatitis (NASH), fibrosis, and cirrhosis and can be complicated by hepatocellular carcinoma. It is strongly associated with metabolic syndromes, obesity, and type 2 diabetes, and it has been shown that metabolic dysregulation is central to its pathogenesis. Recently, it has been suggested that metabolic- (dysfunction) associated fatty liver disease (MAFLD) is a more appropriate term to describe the disease than NAFLD, which puts increased emphasis on the important role of metabolic dysfunction in its pathogenesis. There is strong evidence that mitochondrial dysfunction plays a significant role in the development and progression of NAFLD. Impaired mitochondrial fatty acid oxidation and, more recently, a reduction in mitochondrial quality, have been suggested to play a major role in NAFLD development and progression. In this review, we provide an overview of our current understanding of NAFLD and highlight how mitochondrial dysfunction contributes to its pathogenesis in both animal models and human subjects. Further we discuss evidence that the modification of mitochondrial function modulates NAFLD and that targeting mitochondria is a promising new avenue for drug development to treat NAFLD/NASH.     

Protection of titanium metal by nanohydroxyapatite coating with zirconia and alumina second phases

In this study hydroxyapatite (HA)/zirconia/alumina composite coatings on titanium metal was carried out using Sol-Gel dip coating and calcination process. Hydroxyapatite-Alumina-Zirconia sol, coated samples in three processes by changing final sol stirring time, aging time, calcination temperature of synthesized powder and prepared coating and rate of coating. Some parts of prepared sol were also synthesized and became powder in all three processes. Scanning electron microscopy was used to estimate the particle size of the surface and for morphological analysis. The functional group and crystallization characteristics of the powders were analyzed using (FTIR) and X-Ray diffraction (XRD). Results show that the morphology of HA-Alumina-Zirconia coatings is more homogenous in the second process with 2 hours final sol stirring time, 20 hours aging time under stirring at 60, 675°C calcination temperature for coating and 850°C for powder and 60mm/min rate of dip coating.     

Reactions with 2,3,4,5-Tetrahydro-benzo[b]thiepin-5-one and its Derivatives

2,3,4,5-Tetrahydro-benzo[b]thiepin-5-one, its 7-methyl derivative, and their S, S-dioxides condense with aromatic aldehydes to give the corresponding 4-arylidene derivatives 3 and 4 . Compounds 4 react with hydrazine and phenylhydrazine to yield 3, 3a, 4, 5-tetrahydro-2 H-benzo-[b]thiepino[4, 5-c]pyrazole S, S-dioxide derivatives 5 and 6 . Some of the arylidene derivatives 4 add GRIGNARD reagents to give 7 . The IR spectra of representative examples are discussed.     

Comparing the Effects of Microwave and Conventional Heating Methods on the Evaporation Rate and Quality Attributes of Pomegranate (<Emphasis Type="Italic">Punica granatum</Emphasis> L.) Juice Concentrate

Pomegranate juice was concentrated by conventional heating and microwave heating at different operational pressures (12, 38.5, and 100 kPa), and their effects on evaporation rate and quality attributes of concentrated juice were investigated. The final juice concentration of 40° Brix was achieved in 140, 127, and 109 min at 100, 38.5, and 12 kPa, respectively, by using conventional heating. Applying microwave energy decreased required times to 118, 95, and 75 min. The changes in color, anthocyanin content, and antioxidant capacity during concentration processes were investigated. L*, a*, and b* parameters were measured to estimate the intensity of color loss. All Hunter color parameters decreased with time. Results showed that the degradation of color, anthocyanins, and antioxidant activity were more important in conventional heating compared to microwave heating method. Degradation rates increases by increasing process pressure. A first-order kinetics model was applied to modeling changes in total solid content, anthocyanin content, and antioxidant capacity.     

Exposure of Mice to Thirdhand Smoke Modulates In Vitro and In Vivo Platelet Responses

Smoking is a risk factor for a variety of deleterious conditions, such as cancer, respiratory disease and cardiovascular disease. Thrombosis is an important and common aspect of several cardiovascular disease states, whose risk is known to be increased by both first- and secondhand smoke. More recently, the residual cigarette smoke that persists after someone has smoked (referred to as thirdhand smoke or THS) has been gaining more attention, since it has been shown that it also negatively affects health. Indeed, we have previously shown that 6-month exposure to THS increases the risk of thrombogenesis. However, neither the time-dependence of THS-induced thrombus formation, nor its sex dependence have been investigated. Thus, in the present study, we investigated these issues in the context of a shorter exposure to THS, specifically 3 months, in male and female mice. We show that the platelets from 3-month THS-exposed mice exhibited enhanced activation by agonists. Moreover, we also show that mice of both sexes exposed to THS have decreased tail bleeding as well as decreased thrombus occlusion time. In terms of the role of sex, intersex disparities in thrombus development and hemostasis as well as in platelet aggregation were, interestingly, observed. Together, our findings show that exposing mice to THS for 3 months is sufficient to predispose them to thrombosis; which seems to be driven, at least in part, by an increased activity in platelets, and that it does not manifest equally in both sexes.     

SURFACE ROUGHNESS OF SPUTTERED SILICON. II. MODEL VERIFICATION

Experimental verification of the mathematical surface roughness model for sputtered silicon was performed. The beam shape and its significant level of intensity were determined first by measuring the topography of craters sputtered by focused ion beam (FIB). Then the beam function was generated for various combinations of beam parameters. The material function was developed both by theoretical and experimental analysis. These two functions were then used in the model to calculate the theoretical surface roughness. Microsurface analysis was formed by FIB sputtering of a (100) silicon wafer. The surface roughness at the bottom of the sputtered features was then measured using an atomic force microscope. The theoretical surface roughness was found to be within ±1 and ±5 nm of the measured surface roughness with the measurement uncertainty (standard deviation) of about ±0.36 and ±0.85 nm for R_a and R_t, respectively.