Exploration of the TRIM fold of MuRF1 using EPR reveals a canonical antiparallel structure and extended COS-box

MuRF1 (TRIM63) is a RING-type E3 ubiquitin ligase with a predicted tripartite TRIM fold. TRIM proteins rely upon the correct placement of an N-terminal RING domain, with respect to C-terminal, specific substrate-binding domains. The TRIM domain organization is orchestrated by a central helical domain that forms an antiparallel coiled-coil motif and mediates the dimerization of the fold. MuRF1 has a reduced TRIM composition characterized by a lack of specific substrate binding domains, but contains in its helical domain a conserved sequence motif termed COS-box that has been speculated to fold independently into an α-hairpin. These characteristics had led to question whether MuRF1 adopts a canonical TRIM fold. Using a combination of electron paramagnetic resonance, on spin-labeled protein, and disulfide crosslinking, we show that TRIM63 follows the structural conservation of the TRIM dimerization domain, observed in other proteins. We also show that the COS-box motif folds back onto the dimerization coiled-coil motif, predictably forming a four-helical bundle at the center of the protein and emulating the architecture of canonical TRIMs.publishe

Plasmodium falciparum: linkage disequilibrium between loci in chromosomes 7 and 5 and chloroquine selective pressure in Northern Nigeria.

In view of the recent discovery (Molecular Cell 6, 861-871) of a (Lys76Thr) codon change in gene pfcrt on chromosome 7 which determines in vitro chloroquine resistance in Plasmodium falciparum, we have re-examined samples taken before treatment in our study in Zaria, Northern Nigeria (Parasitology, 119, 343-348). Drug resistance was present in 5/5 cases where the pfcrt 76Thr codon change was seen (100% positive predictive value). Drug sensitivity was found in 26/28 cases where the change was absent (93% negative predictive value). Allele pfcrt 76Thr showed strong linkage disequilibrium with pfmdr1 Tyr86 on chromosome 5, more complete than that between pfcrt and cg2 alleles situated between recombination cross-over points on chromosome 7. Physical linkage of cg2 with pfcrt may account for linkage disequilibrium between their alleles but in the case of genes pfmdr1 and pfcrt, on different chromosomes, it is likely that this is maintained epistatically through the selective pressure of chloroquine

Common variants of the TCF7L2 gene are associated with increased risk of type 2 diabetes mellitus in a UK-resident South Asian population

Background Recent studies have implicated variants of the transcription factor 7-like 2 (TCF7L2) gene in genetic susceptibility to type 2 diabetes mellitus in several different populations. The aim of this study was to determine whether variants of this gene are also risk factors for type 2 diabetes development in a UK-resident South Asian cohort of Punjabi ancestry. Methods We genotyped four single nucleotide polymorphisms (SNPs) of TCF7L2 (rs7901695, rs7903146, rs11196205 and rs12255372) in 831 subjects with diabetes and 437 control subjects. Results The minor allele of each variant was significantly associated with type 2 diabetes; the greatest risk of developing the disease was conferred by rs7903146, with an allelic odds ratio (OR) of 1.31 (95% CI: 1.11 – 1.56, p = 1.96 × 10-3). For each variant, disease risk associated with homozygosity for the minor allele was greater than that for heterozygotes, with the exception of rs12255372. To determine the effect on the observed associations of including young control subjects in our data set, we reanalysed the data using subsets of the control group defined by different minimum age thresholds. Increasing the minimum age of our control subjects resulted in a corresponding increase in OR for all variants of the gene (p ≤ 1.04 × 10-7). Conclusion Our results support recent findings that TCF7L2 is an important genetic risk factor for the development of type 2 diabetes in multiple ethnic groups

Non-Canonicaly Recruited TCRαβCD8αα IELs Recognize Microbial Antigens

In the gut, various subsets of intraepithelial T cells (IELs) respond to self or non-self-antigens derived from the body, diet, commensal and pathogenic microbiota. Dominant subset of IELs in the small intestine are TCRαβCD8αα+ cells, which are derived from immature thymocytes that express self-reactive TCRs. Although most of TCRαβCD8αα+ IELs are thymus-derived, their repertoire adapts to microbial flora. Here, using high throughput TCR sequencing we examined how clonal diversity of TCRαβCD8αα+ IELs changes upon exposure to commensal-derived antigens. We found that fraction of CD8αα+ IELs and CD4+ T cells express identical αβTCRs and this overlap raised parallel to a surge in the diversity of microbial flora. We also found that an opportunistic pathogen (Staphylococcus aureus) isolated from mouse small intestine specifically activated CD8αα+ IELs and CD4+ derived T cell hybridomas suggesting that some of TCRαβCD8αα+ clones with microbial specificities have extrathymic origin. We also report that CD8ααCD4+ IELs and Foxp3CD4+ T cells from the small intestine shared many αβTCRs, regardless whether the later subset was isolated from Foxp3CNS1 sufficient or Foxp3CNS1 deficient mice that lacks peripherally-derived Tregs. Overall, our results imply that repertoire of TCRαβCD8αα+ in small intestine expends in situ in response to changes in microbial flora

TCF7L2 variant genotypes and type 2 diabetes risk in Brazil: significant association, but not a significant tool for risk stratification in the general population

<p>Abstract</p> <p>Background</p> <p>Genetic polymorphisms of the <it>TCF7L2 </it>gene are strongly associated with large increments in type 2 diabetes risk in different populations worldwide. In this study, we aimed to confirm the effect of the <it>TCF7L2 </it>polymorphism <it>rs7903146 </it>on diabetes risk in a Brazilian population and to assess the use of this genetic marker in improving diabetes risk prediction in the general population.</p> <p>Methods</p> <p>We genotyped the single nucleotide polymorphisms (SNP) rs7903146 of the <it>TCF7L2 </it>gene in 560 patients with known coronary disease enrolled in the MASS II (Medicine, Angioplasty, or Surgery Study) Trial and in 1,449 residents of Vitoria, in Southeast Brazil. The associations of this gene variant to diabetes risk and metabolic characteristics in these two different populations were analyzed. To access the potential benefit of using this marker for diabetes risk prediction in the general population we analyzed the impact of this genetic variant on a validated diabetes risk prediction tool based on clinical characteristics developed for the Brazilian general population.</p> <p>Results</p> <p>SNP rs7903146 of the <it>TCF7L2 </it>gene was significantly associated with type 2 diabetes in the MASS-II population (OR = 1.57 per T allele, p = 0.0032), confirming, in the Brazilian population, previous reports of the literature. Addition of this polymorphism to an established clinical risk prediction score did not increased model accuracy (both area under ROC curve equal to 0.776).</p> <p>Conclusion</p> <p><it>TCF7L2 </it>rs7903146 T allele is associated with a 1.57 increased risk for type 2 diabetes in a Brazilian cohort of patients with known coronary heart disease. However, the inclusion of this polymorphism in a risk prediction tool developed for the general population resulted in no improvement of performance. This is the first study, to our knowledge, that has confirmed this recent association in a South American population and adds to the great consistency of this finding in studies around the world. Finally, confirming the biological association of a genetic marker does not guarantee improvement on already established screening tools based solely on demographic variables.</p

Expression of MuRF1 or MuRF2 is essential for the induction of skeletal muscle atrophy and dysfunction in a murine pulmonary hypertension model

Background Pulmonary hypertension leads to right ventricular heart failure and ultimately to cardiac cachexia. Cardiac cachexia induces skeletal muscles atrophy and contractile dysfunction. MAFbx and MuRF1 are two key proteins that have been implicated in chronic muscle atrophy of several wasting states. Methods Monocrotaline (MCT) was injected over eight weeks into mice to establish pulmonary hypertension as a murine model for cardiac cachexia. The effects on skeletal muscle atrophy, myofiber force, and selected muscle proteins were evaluated in wild-type (WT), MuRF1, and MuRF2-KO mice by determining muscle weights, in vitro muscle force and enzyme activities in soleus and tibialis anterior (TA) muscle. Results In WT, MCT treatment induced wasting of soleus and TA mass, loss of myofiber force, and depletion of citrate synthase (CS), creatine kinase (CK), and malate dehydrogenase (MDH) (all key metabolic enzymes). This suggests that the murine MCT model is useful to mimic peripheral myopathies as found in human cardiac cachexia. In MuRF1 and MuRF2-KO mice, soleus and TA muscles were protected from atrophy, contractile dysfunction, while metabolic enzymes were not lowered in MuRF1 or MuRF2-KO mice. Furthermore, MuRF2 expression was lower in MuRF1KO mice when compared to C57BL/6 mice. Conclusions In addition to MuRF1, inactivation of MuRF2 also provides a potent protection from peripheral myopathy in cardiac cachexia. The protection of metabolic enzymes in both MuRF1KO and MuRF2KO mice as well as the dependence of MuRF2 expression on MuRF1 suggests intimate relationships between MuRF1 and MuRF2 during muscle atrophy signaling

Response-adapted treatment with rituximab, bendamustine, mitoxantrone, and dexamethasone followed by rituximab maintenance in patients with relapsed or refractory follicular lymphoma after first-line immunochemotherapy: Results of the RBMDGELTAMO08 phase II trial

Background Consensus is lacking regarding the optimal salvage therapy for patients with follicular lymphoma who relapse after or are refractory to immunochemotherapy. Methods This phase II trial evaluated the efficacy and safety of response-adapted therapy with rituximab, bendamustine, mitoxantrone, and dexamethasone (RBMD) in follicular lymphoma patients who relapsed after or were refractory to first-line immunochemotherapy. Sixty patients received three treatment cycles, and depending on their response received an additional one (complete/unconfirmed complete response) or three (partial response) cycles. Patients who responded to induction received rituximab maintenance therapy for 2 years. Results Thirty-three (55%) and 42 (70%) patients achieved complete/unconfirmed complete response after three cycles and on completing induction therapy (4-6 cycles), respectively (final overall response rate, 88.3%). Median progression-free survival was 56.4 months (median follow-up, 28.3 months; 95% CI, 15.6-51.2). Overall survival was not reached. Progression-free survival did not differ between patients who received four vs six cycles (P = .6665), nor between patients who did/did not receive rituximab maintenance after first-line therapy (P = .5790). Median progression-free survival in the 10 refractory patients was 25.5 months (95% CI, 0.6-N/A) and was longer in patients who had shown progression of disease after 24 months of first-line therapy (median, 56.4 months; 95% CI, 19.8-56.4) than in those who showed early progression (median, 42.31 months; 95% CI, 24.41-NA) (P = .4258). Thirty-six (60%) patients had grade 3/4 neutropenia. Grade 3/4 febrile neutropenia and infection were recorded during induction (4/60 [6.7%] and 5/60 [8.3%] patients, respectively) and maintenance (2/43 [4.5%] and 4/43 [9.1%] patients, respectively). Conclusions This response-adapted treatment with RBMD followed by rituximab maintenance is an effective and well-tolerated salvage treatment for relapsed/refractory follicular lymphoma following first-line immunochemotherapy

Weak or no association of TCF7L2 variants with Type 2 diabetes risk in an Arab population

<p>Abstract</p> <p>Background</p> <p>The rs7903146 and rs12255372 variants of <it>TCF7L2 </it>have been strongly associated with type 2 diabetes (T2D) risk in most populations studied to date. Meta-analysis of 27 different studies has resulted in a global OR of 1.46 [1.42–1.51] (rs7903146 variant). Thus far, despite a high incidence of T2D, the role of this variant in Arabs has not been established.</p> <p>Methods</p> <p>We performed a case-control association study using 522 Saudi T2D patients (WHO criteria), and 346 controls (age > 60; fasting plasma glucose < 7 mmol/L). Genotyping was performed by pyrosequencing. Statistical analyses were performed using SPSS version 13.0 for Windows (SPSS, Chicago, IL, USA).</p> <p>Results</p> <p>For rs7903146, the T allele frequency of the cases (0.415) was not different from that observed in the controls (0.405). The crude odds ratio (OR) was 1.04 with a 95% CI of 0.86–1.27 (P = 0.675). For rs12255372, the T allele frequency of the cases (0.368) was not different from that observed in the controls (0.355). Retrospective power calculations based upon an OR of 1.46 reported in a comprehensive meta-analysis of <it>TCF7L2 </it>risk, indicated this study was sufficiently powered (96.92%; α = 0.05) to detect an effect of similar magnitude to that reported for rs7903146.</p> <p>Conclusion</p> <p>Our study is consistent with weak or no association of T2D in Arabs with the two <it>TCF7L2 </it>variants, however it cannot rule out an effect of other SNPs in this gene. Future studies in this population are required to confirm our findings and may indicate the presence of yet to be defined genetic risk factors for T2D.</p

Variations in Suppressor Molecule CTLA-4 Gene Are Related to Susceptibility to Multiple Myeloma in a Polish Population

Various phenotype and functional T-cell abnormalities are observed in multiple myeloma (MM) patients. The aim of this study was to investigate the association between polymorphisms in the gene encoding cytotoxic T-lymphocyte antigen-4 (CTLA-4), a negative regulator of the T-lymphocyte immune response and susceptibility to multiple myeloma in a Polish population. Two hundred MM patients and 380 healthy subjects were genotyped for the following polymorphisms: CTLA-4c.49A>G, CTLA-4g.319C>T, CTLA-4g.*642AT(8_33), CT60 (CTLA-4g.*6230G>A), Jo31 (CTLA-4g.*10223G>T). Our study is the largest and most comprehensive evaluation to date of the association between genetic polymorphisms in the CTLA-4 molecule and multiple myeloma. It was found that CTLA-4c.49A>G[G], CT60[G], and Jo31[G] alleles were more frequently observed in MM patients than in controls (0.50 vs. 0.44, p = 0.03, 0.65 vs. 0.58, p = 0.04, and 0.63 vs. 0.57, p = 0.03, respectively). Moreover, the haplotype CTLA-4c.49A>G[G], CTLA-4g.319C>T[C], CTLA-4g.*642AT(8_33) [8], CT60[G], Jo31[G] including all susceptibility alleles increases the risk of MM about fourfold (OR: 3.79, 95%CI: 2.08–6.89, p = 0.00001). These findings indicate that genetic variations in the CTLA-4 gene play role in susceptibility to multiple myeloma and warrant further investigation through replication studies