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The virulence factor mycolactone is responsible for the immunosuppression and tissue necrosis that characterise Buruli ulcer, a disease caused by infection with Mycobacterium ulcerans. In this study, we confirm that Sec61, the protein-conducting channel that coordinates entry of secretory proteins into the endoplasmic reticulum, is a primary target of mycolactone, and characterise the nature of its inhibitory effect. We conclude that mycolactone constrains the ribosome-nascent chain-Sec61 complex, consistent with its broad-ranging perturbation of the co-translational translocation of classical secretory proteins. In contrast, the effect of mycolactone on the post-translational, ribosome-independent translocation of short secretory proteins through the Sec61 complex is dependent on both signal sequence hydrophobicity and the translocation competence of the mature domain. Changes to protease sensitivity strongly suggest that mycolactone acts by inducing a conformational change in the pore-forming Sec61α subunit. These findings establish that mycolactone inhibits Sec61-mediated protein translocation and highlight differences between the co- and post-translational routes that the Sec61 complex mediates. We propose that mycolactone also provides a useful tool for further delineating the molecular mechanisms of Sec61-dependent protein translocation.

Rpn13 is an intrinsic ubiquitin receptor of the 26S proteasome regulatory subunit that facilitates substrate capture prior to degradation. Here we show that the C-terminal region of Rpn13 binds to the tetratricopeptide repeat (TPR) domain of SGTA, a cytosolic factor implicated in the quality control of mislocalised membrane proteins (MLPs). The overexpression of SGTA results in a substantial increase in steady-state MLP levels, consistent with an effect on proteasomal degradation. However, this effect is strongly dependent upon the interaction of SGTA with the proteasomal component Rpn13. Hence, overexpression of the SGTA-binding region of Rpn13 or point mutations within the SGTA TPR domain both inhibit SGTA binding to the proteasome and substantially reduce MLP levels. These findings suggest that SGTA can regulate the access of MLPs to the proteolytic core of the proteasome, implying that a protein quality control cycle that involves SGTA and the BAG6 complex can operate at the 19S regulatory particle. We speculate that the binding of SGTA to Rpn13 enables specific polypeptides to escape proteasomal degradation and/or selectively modulates substrate degradation., Highlighted Article: Binding of SGTA to the proteasome delays substrate degradation, thereby providing a mechanism for potentially viable proteins to be rescued for reuse.

We used a collection of 708 prospectively collected autopsied brains to assess the methylation state of the brain's DNA in relation to Alzheimer's disease (AD). We found that the level of methylation at 71 of the 415,848 interrogated CpGs was significantly associated with the burden of AD pathology, including CpGs in the ABCA7 and BIN1 regions, which harbor known AD susceptibility variants. We validated 11 of the differentially methylated regions in an independent set of 117 subjects. Furthermore, we functionally validated these CpG associations and identified the nearby genes whose RNA expression was altered in AD: ANK1, CDH23, DIP2A, RHBDF2, RPL13, SERPINF1 and SERPINF2. Our analyses suggest that these DNA methylation changes may have a role in the onset of AD given that we observed them in presymptomatic subjects and that six of the validated genes connect to a known AD susceptibility gene network., National Institutes of Health (U.S.) (Grant R01 AG036042), National Institutes of Health (U.S.) (Grant R01AG036836), National Institutes of Health (U.S.) (Grant R01 AG17917), National Institutes of Health (U.S.) (Grant R01AG15819), National Institutes of Health (U.S.) (Grant R01 AG032990), National Institutes of Health (U.S.) (Grant R01 AG18023), National Institutes of Health (U.S.) (Grant RC2 AG036547), National Institutes of Health (U.S.) (Grant P30 AG10161), National Institutes of Health (U.S.) (Grant P50 AG016574), National Institutes of Health (U.S.) (Grant U01 ES017155), National Institutes of Health (U.S.) (Grant KL2 RR024151), National Institutes of Health (U.S.) (Grant K25 AG041906-01)

We present the first identification of transient folding intermediates of endogenous thyroglobulin (Tg; a large homodimeric secretory glycoprotein of thyrocytes), which include mixed disulfides with endogenous oxidoreductases servicing Tg folding needs. Formation of disulfide-linked Tg adducts with endoplasmic reticulum (ER) oxidoreductases begins cotranslationally. Inhibition of ER glucosidase activity blocked formation of a subgroup of Tg adducts containing ERp57 while causing increased Tg adduct formation with protein disulfide isomerase (PDI), delayed adduct resolution, perturbed oxidative folding of Tg monomers, impaired Tg dimerization, increased Tg association with BiP/GRP78 and GRP94, activation of the unfolded protein response, increased ER-associated degradation of a subpopulation of Tg, partial Tg escape from ER quality control with increased secretion of free monomers, and decreased overall Tg secretion. These data point towards mixed disulfides with the ERp57 oxidoreductase in conjunction with calreticulin/calnexin chaperones acting as normal early Tg folding intermediates that can be "substituted" by PDI adducts only at the expense of lower folding efficiency with resultant ER stress.

Existing efficacy trials of Omega-3 (omega-3) fatty acids in mood disorders have yielded inconsistent results. The current paper is an effort to provide a systematic review and meta-analysis to evaluate efficacy of omega-3 fatty acids in treatment of mood disorders.We searched Medline, Embase, PsychInfo, and the Cochrane Controlled Trials registry up to June 2008 for randomized trials investigating efficacy of omega-3 fatty acids in mood disorders.We conducted random effects meta-analyses.We used the I2 statistic to quantify between-study inconsistency, and conducted pre-specified subgroup analyses to explore potential explanations for inconsistency.We included 21 trials in our systematic review and found 13 trials to be eligible for meta-analysis. The pooled standardized mean difference in depressed mood states (n = 554 in 12 trials) was -0.47 (95% CI:-0.92,-0.02; I2 = 82.7; p = 0.07) and in manic mood states (n = 126 in 4 trials) was 0.22 (95% CI: -0.21, 0.65; I2 = 40.5; p = 0.31).We did not identify any treatment- subgroup interaction across forms of omega-3 fatty acids preparations (P = 0.99) or patient diagnosis (bipolar vs. unipolar depressive disorder; P = 0.96); there was a significant correlation between omega-3 fatty acids dose and treatment effect on depressive symptoms (r = 0.5, p = 0.04), but not on manic symptoms (P = 0.3).The available evidence suggests that omega-3 fatty acids are a potential treatment of depressive disorders, but not mania. The unexplained between-study inconsistency and imprecision of the pooled estimates mitigate this suggestion. Large randomized placebo-controlled trials are needed to better estimate the value of this intervention for patients with depression.

Two distinct protein targeting pathways can direct proteins to the Escherichia coli inner membrane. The Sec pathway involves the cytosolic chaperone SecB that binds to the mature region of pre-proteins. SecB targets the pre-protein to SecA that mediates preprotein translocation through the SecYEG translocon. The SRP pathway is probably used primarily for the targeting and assembly of inner membrane proteins. It involves the signal recognition particle (SRP) that interacts with the hydrophobic targeting signal of nascent proteins. By using a protein cross-linking approach, we demonstrate here that the SRP pathway delivers nascent inner membrane proteins at the membrane. The SRP receptor FtsY, GTP and inner membranes are required for release of the nascent proteins from the SRP. Upon release of the SRP at the membrane, the targeted nascent proteins insert into a translocon that contains at least SecA, SecY and SecG. Hence, as appears to be the case for several other translocation systems, multiple targeting mechanisms deliver a variety of precursor proteins to a common membrane translocation complex of the E.coli inner membrane.