Chronic ethanol consumption is known as an independent risk factor for type 2 diabetes, which is characterized by impaired glucose homeostasis and insulin resistance; however, there is a great deal of controversy concerning the relationships between alcohol consumption and the development of type 2 diabetes. We investigated the effects of chronic ethanol consumption on pancreatic ␤-cell dysfunction and whether generated peroxynitrite participates in the impaired glucose homeostasis. Here we show that chronic ethanol feeding decreases the ability of pancreatic ␤-cells to mediate insulin secretion and ATP production in coordination with the decrease of glucokinase, Glut2, and insulin expression. Specific blockade of ATF3 using siRNA or C-terminally deleted ATF3(⌬C) attenuated ethanol-induced pancreatic ␤-cell apoptosis or dysfunction and restored the down-regulation of glucokinase (GCK), insulin, and pancreatic duodenal homeobox-1 induced by ethanol. GCK inactivation and down-regulation were predominantly mediated by ethanol metabolism-generated peroxynitrite, which were suppressed by the peroxynitrite scavengers N ␥ -monomethyl-L-arginine, uric acid, and deferoxamine but not by the S-nitrosylation inhibitor DTT, indicating that tyrosine nitration is the predominant modification associated with GCK down-regulation and inactivation rather than S-nitrosylation of cysteine. Tyrosine nitration of GCK prevented its association with pBad, and GCK translocation into the mitochondria results in subsequent proteasomal degradation of GCK following ubiquitination. This study identified a novel and efficient pathway by which chronic ethanol consumption may induce GCK down-regulation and inactivation by inducing tyrosine nitration of GCK, resulting in pancreatic ␤-cell apoptosis and dysfunction. Peroxynitrite-induced ATF3 may also serve as a potent upstream regulator of GCK down-regulation and ␤-cell apoptosis.

Epidemiological and phylogenetic studies of hepatitis C virus (HCV) have identified six major HCV genotypes and have attempted to characterize their origin and spread worldwide. Putative regions of endemic infection have been identified for all HCV genotypes except HCV genotype 5a. Although HCV genotype 5a was previously thought to be largely restricted to the northern part of South Africa, this study reports an unexpected cluster of the genotype in West Flanders Province in Belgium. To investigate the molecular epidemiology of this cluster and of HCV genotype 5a in general, a rigorous phylogenetic analysis of Belgian and South African HCV genotype 5a samples was performed. Remarkably, the Belgian and South African strains form two distinct clusters of similar diversity. We used a Bayesian coalescent method to estimate the rate of virus spread through time for HCV genotype 5a in both regions. Our results indicate that HCV genotype 5a strains have been spreading independently in Belgium and South Africa for more than 100 years, with a rate of spread characteristic of an epidemic genotype. These findings have major implications for tracing the origin of HCV genotype 5a. Here, we speculate about the possible origins of these clusters.With an estimated 170 million people infected worldwide, hepatitis C virus (HCV) is responsible for most cases of parenteral non-A, non-B hepatitis and is therefore the leading cause of chronic liver disease and hepatocellular carcinoma (38). The virus is responsible for 10,000 deaths per year in the United States alone, and this number is expected to increase substantially in the forthcoming decades (1). HCV is a bloodborne pathogen; risk factors for infection include injecting drug use, receipt of a blood transfusion before 1990, use of inadequately sterilized medical equipment, and use of scarification and tattooing tools (14). Sexual and perinatal transmissions rarely occur, except in human immunodeficiency virus-HCV-coinfected individuals (4). A nonnegligible proportion (about 20%) of HCV infections have an "undefined" route of transmission.HCV is a small, enveloped human virus containing a singlestranded RNA genome of about 9,600 nucleotides (nt) (36). HCV displays a high degree of genetic variability (22) and is classified into six major genotypes that show sequence similarities of only 66% to 69%. Each genotype contains multiple subtypes with Ͼ75% nucleotide sequence similarity (29). In comparison, the degree of similarity among different virions in a single infected patient is Ͼ95% (11).Importantly, numerous studies have revealed a relationship between the HCV genotype and the response to interferon or pegylated interferon therapy, alone or in combination with ribavirin. Patients infected with HCV genotype 1 respond less to therapy, while patients infected with HCV type 2 or 3 show the best response (23). Since HCV genotype 5a infections are scarce, treatment responses for HCV genotype 5a are largely unknown (15). For HCV genotype 6a infections, treatment response data are als...

RhoA is overexpressed in human cancer and contributes to aberrant cell motility and metastatic progression; however, regulatory mechanisms controlling RhoA activity in cancer are poorly understood. Neuroepithelial transforming gene 1 (Net1) is a RhoA guanine nucleotide exchange factor that is overexpressed in human cancer. It encodes two isoforms, Net1 and Net1A, which cycle between the nucleus and plasma membrane. Net1 proteins must leave the nucleus to activate RhoA, but mechanisms controlling the extranuclear localization of Net1 isoforms have not been described. Here, we show that Rac1 activation causes relocalization of Net1 isoforms outside the nucleus and stimulates Net1A catalytic activity. These effects do not require Net1A catalytic activity, its pleckstrin homology domain, or its regulatory C terminus. We also show that Rac1 activation protects Net1A from proteasome-mediated degradation. Replating cells on collagen stimulates endogenous Rac1 to relocalize Net1A, and inhibition of proteasome activity extends the duration and magnitude of Net1A relocalization. Importantly, we demonstrate that Net1A, but not Net1, is required for cell spreading on collagen, myosin light chain phosphorylation, and focal adhesion maturation. These data identify the first physiological mechanism controlling the extranuclear localization of Net1 isoforms. They also demonstrate a previously unrecognized role for Net1A in regulating cell adhesion.

This was an exploratory study to investigate lamivudine-resistant hepatitis B virus (HBV) strains in selected lamivudine-naïve HBV carriers with and without human immunodeficiency virus (HIV) co-infection in South African patients. Thirty-five lamivudine-naïve HBV infected patients with or without HIV co-infection were studied: 15 chronic HBV mono-infected patients and 20 HBV-HIV co-infected patients. The latter group was further sub-divided into 13 occult HBV (HBsAg-negative) and 7 overt HBV (HBsAg- positive) patients. HBsAg, anti-HBs, anti-HBc, and anti-HIV 1/2 were determined as part of routine diagnosis using Axsym assays (Abbott Laboratories, North Chicago, IL). Serum samples were PCR amplified with HBV reverse transcriptase (RT) primers, followed by direct sequencing across the tyrosine-methionine-aspartate-aspartate (YMDD) motif of the major catalytic region in the C domain of the HBV RT enzyme. HBV viral load was performed with Amplicor HBV Monitor test v2.0 (Roche Diagnostics, Penzberg, Germany). HBV lamivudine-resistant strains were detected in 3 of 15 mono-infected chronic hepatitis B patients and 10 of 20 HBV-HIV co-infected patients. To the best of our knowledge, this constitutes the first report of HBV lamivudine-resistant strains in therapy-naïve HBV-HIV co-infected patients. The HBV viral loads for mono-infected and co-infected patients ranged from 3.32 x 10(2) to 3.82 x 10(7) and <200 to 4.40 x 10(3) copies/ml, respectively. It remains to be seen whether such pre-existing antiviral mutations could result in widespread emergence of HBV resistant strains when lamivudine-containing highly active antiretroviral (ARV) treatment (HAART) regimens become widely applied in South Africa, as this is likely to have potential implications in the management of HBV-HIV co-infected patients.