Neurodegenerative diseases are characterized by the accumulation of misfolded proteins in the brain. Insights into protein quality control mechanisms to prevent neuronal dysfunction and cell death ...are crucial in developing causal therapies. Here, we report that various disease‐associated protein aggregates are modified by the linear ubiquitin chain assembly complex (LUBAC). HOIP, the catalytic component of LUBAC, is recruited to misfolded Huntingtin in a p97/VCP‐dependent manner, resulting in the assembly of linear polyubiquitin. As a consequence, the interactive surface of misfolded Huntingtin species is shielded from unwanted interactions, for example with the low complexity sequence domain‐containing transcription factor Sp1, and proteasomal degradation of misfolded Huntingtin is facilitated. Notably, all three core LUBAC components are transcriptionally regulated by Sp1, linking defective LUBAC expression to Huntington's disease. In support of a protective activity of linear ubiquitination, silencing of OTULIN, a deubiquitinase with unique specificity for linear polyubiquitin, decreases proteotoxicity, whereas silencing of HOIP has the opposite effect. These findings identify linear ubiquitination as a protein quality control mechanism and hence a novel target for disease‐modifying strategies in proteinopathies.
Synopsis
Misfolded proteins recruit the linear ubiquitin chain assembly complex (LUBAC) via p97/VCP, resulting in the assembly of linear ubiquitin chains at protein aggregates, such as Huntingtin with an polyQ expansion (Htt‐polyQ). This modification interferes with proteotoxicity of Htt‐polyQ by preventing sequestration of the transcription factor Sp1 and by promoting proteasomal degradation of misfolded Htt‐polyQ.
The linear ubiquitin chain assembly complex (LUBAC) is recruited to misfolded protein species, such as mutant Huntingtin, Ataxin‐3, SOD1, and TDP‐43.
Linear ubiquitination of mutant Huntingtin causes remodeling of its interactive surface.
Expression of LUBAC is dysregulated in Huntington's disease by sequestration of the transcription factor Sp1 to misfolded Huntingtin.
Linear ubiquitination promotes removal of misfolded protein species in a p97/VCP‐ and proteasome‐dependent manner.
Recruitment of the LUBAC E3 ligase, best known for its function in NF‐κB signalling, to protein aggregates for linear ubiquitin‐mediated removal of misfolded neurodegenerative disease proteins such as huntingtin, SOD1 or TDP‐43 exemplifies a new, potentially targetable cellular mechanism for controlling proteinopathies.
c-MYC is an oncogenic transcription factor that is degraded by the proteasome pathway. However, the mechanism that regulates delivery of c-MYC to the proteasome for degradation is not well ...characterized. Here, the results show that the motor protein complex Kinesin-1 transports c-MYC to the cytoplasm for proteasomal degradation. Inhibition of Kinesin-1 function enhanced ubiquitination of c-MYC and induced aggregation of c-MYC in the cytoplasm. Transport studies showed that the c-MYC aggregates moved from the nucleus to the cytoplasm and KIF5B is responsible for the transport in the cytoplasm. Furthermore, inhibition of the proteasomal degradation process also resulted in an accumulation of c-MYC aggregates in the cytoplasm. Moreover, Kinesin-1 was shown to interact with c-MYC and the proteasome subunit S6a. Inhibition of Kinesin-1 function also reduced c-MYC-dependent transformation activities. Taken together, the results strongly suggest that Kinesin-1 transports c-MYC for proteasomal degradation in the cytoplasm and the proper degradation of c-MYC mediated by Kinesin-1 transport is important for transformation activities of c-MYC. In addition, the results indicate that Kinesin-1 transport mechanism is important for degradation of a number of other proteins as well.
•Kinesin-1 transports c-MYC to the cytoplasm for proteasomal degradation.•Inhibition of Kinesin-1 enhances ubiquitination and aggregation of c-MYC.•Reduction of proteasomal degradation leads to accumulation of c-MYC aggregates.•Kinesin-1 interacts with c-MYC and the proteasome subunit S6a.•Inhibition of Kinesin-1 reduces c-MYC-dependent transformation activities.
The emergence and spread of cryptococcosis caused by the Cryptococcus gattii species complex has become a major public concern worldwide. C. deuterogattii (VGIIa) outbreaks in the Pacific Northwest ...region demonstrate the expansion of this fungal infection to temperate climate regions. However, infections due to the C. gattii species complex in China have rarely been reported. In this study, we studied eleven clinical strains of the C. gattii species complex isolated from Guangxi, southern China. The genetic identity and variability of these isolates were analyzed via multi-locus sequence typing (MLST), and the phylogenetic relationships among these isolates and global isolates were evaluated. The mating type, physiological features and antifungal susceptibilities of these isolates were also characterized. Among the eleven isolates, six belonged to C. deuterogattii, while five belonged to C. gattii sensu stricto. The C. deuterogattii strains from Guangxi, southern China were genetically variable and clustered with different clinical isolates from Brazil. All strains were MATalpha, and three C. deuterogattii isolates (GX0104, GX0105 and GX0147) were able to undergo sexual reproduction. Moreover, most strains had capsule and were capable of melanin production when compared to the outbreak strain from Canada. Most isolates were susceptible to antifungal drugs; yet one of eleven immunocompetent patients died of cryptococcal meningitis caused by C. deuterogattii (GX0147). Our study indicated that the highly pathogenic C. deuterogattii may be emerging in southern China, and effective nationwide surveillance of C. gattii species complex infection is necessary.
Approximately 50% of glioblastomas (GBMs) are characterized by overexpression of the epidermal growth factor receptor (EGFR) and EGFR gene amplification. In approximately 25% of instances, ...constitutively activated EGFR mutants are present. These observations make EGFR-inhibiting drugs a logical approach for trials in recurrent GBM.
In a randomized, controlled, phase II trial, 110 patients with progressive GBM after prior radiotherapy were randomly assigned to either erlotinib or a control arm that received treatment with either temozolomide or carmustine (BCNU). The primary end point was 6-month progression-free survival (PFS). Tumor specimens obtained at first surgery were investigated for EGFR expression; EGFRvIII mutants; EGFR amplification; EGFR mutations in exons 18, 19, and 21; and pAkt. These results were correlated with outcome. Pharmacokinetic analysis was part of the study. RESULTS; Treatment was well tolerated in general; skin toxicity was the most frequent adverse effect of erlotinib. The 6-month PFS rate in the erlotinib arm was 11.4% (95% CI, 4.6% to 21.5%), and it was 24% in the control arm. Of all explored biomarkers, only low pAkt expression appeared to be of borderline significance to an improved outcome. None of the eight patients who had tumors with EGFRvIII mutant presence and PTEN expression had 6-month PFS. The use of enzyme-inducing anticonvulsants significantly increased erlotinib clearance, but pharmacokinetic findings were not related to outcome.
Erlotinib has insufficient single-agent activity in unselected GBM. No clear biomarker associated with improved outcome to erlotinib was identified.
The present study aimed at the molecular characterization of pathogenic and non pathogenic F. oxysporum f. sp. lycopersici strains isolated from tomato. The causal agent isolated from symptomatic ...plants and soil samples was identified based on morphological and molecular analyses. Pathogenicity testing of 69 strains on five susceptible tomato varieties showed 45% of the strains were highly virulent and 30% were moderately virulent. Molecular analysis based on the fingerprints obtained through ISSR indicated the presence of wide genetic diversity among the strains. Phylogenetic analysis based on ITS sequences showed the presence of at least four evolutionary lineages of the pathogen. The clustering of F. oxysporum with non pathogenic isolates and with the members of other formae speciales indicated polyphyletic origin of F. oxysporum f. sp. lycopersici. Further analysis revealed intraspecies variability and nucleotide insertions or deletions in the ITS region among the strains in the study and the observed variations were found to be clade specific. The high genetic diversity in the pathogen population demands for development of effective resistance breeding programs in tomato. Among the pathogenic strains tested, toxigenic strains harbored the Fum1 gene clearly indicating that the strains infecting tomato crops have the potential to produce Fumonisin.
Peripheral arterial disease (PAD) is the least studied complication of nephrotic syndrome (NS). Risk factors which predispose children with NS to developing PAD include hyperlipidaemia, hypertension ...and prolonged use of steroids. The development of PAD significantly increases the morbidity and mortality associated with NS as such children are prone to sudden cardiac death. The ankle brachial index (ABI) is a tool that has been proven to have high specificity and sensitivity in detecting PAD even in asymptomatic individuals. We aimed to determine the prevalence of PAD in children with NS and to identify risk factors that can independently predict its development. A comparative cross-sectional study was conducted involving 200 subjects (100 with NS and 100 apparently healthy comparative subjects that were matched for age, sex and socioeconomic class). Systolic blood pressures were measured in all limbs using the pocket Doppler machine (Norton Doppler scan machine). ABI was calculated as a ratio of ankle to arm systolic blood pressure. PAD was defined as ABI less than 0.9. The prevalence of PAD was significantly higher in children with NS than matched comparison group (44.0% vs 6.0%, p < 0.001). Average values of waist and hip circumference were significantly higher in subjects with PAD than those without PAD (61.68± 9.1cm and 67.6± 11.2 cm vs 57.03 ± 8.3cm and 65.60± 12.5cm respectively, p< 0.005). Serum lipids (triglyceride, very low density lipoprotein, total cholesterol and low density lipoprotein) were also significantly higher in subjects with PAD than those without PAD 106.65mg/dl (67.8–136.7) vs 45.72mg/dl (37.7–61.3), 21.33mg/dl (13.6–27.3) vs 9.14mg/dl (7.5–12.3), 164.43mg/dl (136.1–259.6) vs 120.72mg/dl (111.1–142.1) and 93.29mg/dl (63.5–157.3) vs 61.84mg/dl (32.6–83.1), respectively p< 0.05. Increasing duration since diagnosis of NS, having a steroid resistant NS and increasing cumulative steroid dose were independent predictors of PAD in children with NS; p< 0.05 respectively. With these findings, it is recommended that screening for PAD in children with NS should be done to prevent cardiovascular complications before they arise.
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•In-liquid sensitivity of shear SMRs can be boosted using specific designs.•This enhancement can be achieved without increasing the operating frequency.•Low impedance electrodes ...increase the sensitivity to density and viscosity.•Energy trapping effects at the sensing interface makes this possible.•The same effect occurs when applied as in liquid gravimetric sensors.
In contrast to quartz crystal microbalances, the sensitivity of thin film bulk acoustic wave resonators (FBARs) is strongly dependent on all films comprising the multilayered structure. Previous studies proved that placing low acoustic impedance materials at the sensing surface of longitudinal-mode FBARs operating in air can enhance their mass sensitivity by modifying the energy trapped at the sensing surface. Here we investigate if the in-liquid sensitivity to density-viscosity and mass of shear-mode AlN-based solidly mounted resonators displays a similar dependence on the device configuration. By using the finite element simulation method accompanied by experimental verifications we demonstrate that for a given value of the resonant frequency, the sensitivity can be boosted by a proper design of the devices. The results can be of application to in-liquid physical sensors or to gravimetric chemical or biological sensors.
Estrogens and androgens have both been implicated as causes of benign prostatic hyperplasia (BPH). Although epidemiological data on an association between serum androgen concentrations and BPH are ...inconsistent, it is generally accepted that androgens play a permissive role in BPH pathogenesis. In clinical practice, inhibitors of 5α-reductase (which converts testosterone to the more potent androgen dihydrotestosterone) have proven effective in the management of BPH, confirming an essential role for androgens in BPH pathophysiology. To date, multiple lines of evidence support a role for estrogens in BPH pathogenesis. Studies of the two estrogen receptor (ER) subtypes have shed light on their differential functions in the human prostate; ERα and ERβ have proliferative and antiproliferative effects on prostate cells, respectively. Effects of estrogens on the prostate are associated with multiple mechanisms including apoptosis, aromatase expression and paracrine regulation via prostaglandin E2. Selective estrogen receptor modulators or other agents that can influence intraprostatic estrogen levels might conceivably be potential therapeutic targets for the treatment of BPH.
Here we demonstrate that biomolecular contaminants, such as nucleic acid molecules, can seriously interfere with immobilized metal ion affinity chromatography (IMAC)-based phosphopeptide enrichments. ...We address and largely solve this issue, developing a robust protocol implementing methanol/chloroform protein precipitation and enzymatic digestion using benzonase, which degrades all forms of DNA and RNA, before IMAC-column loading. This simple procedure resulted in a drastic increase of enrichment sensitivity, enabling the identification of around 17,000 unique phosphopeptides and 12,500 unambiguously localized phosphosites in human cell-lines from a single LC-MS/MS run, constituting a 50% increase when compared with the standard protocol. The improved protocol was also applied to bacterial samples, increasing the number of identified bacterial phosphopeptides even more strikingly, by a factor 10, when compared with the standard protocol. For E. coli we detected around 1300 unambiguously localized phosphosites per LC-MS/MS run. The preparation of these ultra-pure phosphopeptide samples only requires marginal extra costs and sample preparation time and should thus be adoptable by every laboratory active in the field of phosphoproteomics.
Many decisions regarding health resource utilization flow through the patient-clinician interaction. Thus, it represents a place where de-implementation interventions may have considerable effect on ...reducing the use of clinical interventions that lack efficacy, have risks that outweigh benefits, or are not cost-effective (i.e., low-value care). The objective of this systematic review with meta-analysis was to determine the effect of de-implementation interventions that engage patients within the patient-clinician interaction on use of low-value care.
MEDLINE, EMBASE, and CINAHL were searched from inception to November 2019. Gray literature was searched using the CADTH tool. Studies were screened independently by two reviewers and were included if they (1) described an intervention that engaged patients in an initiative to reduce low-value care, (2) reported the use of low-value care with and without the intervention, and (3) were randomized clinical trials (RCTs) or quasi-experimental designs. Studies describing interventions solely focused on clinicians or published in a language other than English were excluded. Data was extracted independently in duplicate and pertained to the low-value clinical intervention of interest, components of the strategy for patient engagement, and study outcomes. Quality of included studies was assessed using the Cochrane Risk of Bias tool for RCTs and a modified Downs and Black checklist for quasi-experimental studies. Random effects meta-analysis (reported as risk ratio, RR) was used to examine the effect of de-implementation interventions on the use of low-value care.
From 6736 unique citations, 9 RCTs and 13 quasi-experimental studies were included in the systematic review. Studies mostly originated from the USA (n = 13, 59%), targeted treatments (n = 17, 77%), and took place in primary care (n = 10, 45%). The most common intervention was patient-oriented educational material (n = 18, 82%), followed by tools for shared decision-making (n = 5, 23%). Random effects meta-analysis demonstrated that de-implementation interventions that engage patients within the patient-clinician interaction led to a significant reduction in low-value care in both RCTs (RR 0.74; 95% CI 0.66-0.84) and quasi-experimental studies (RR 0.61; 95% CI 0.43-0.87). There was significant inter-study heterogeneity; however, intervention effects were consistent across subgroups defined by low-value practice and patient-engagement strategy.
De-implementation interventions that engage patients within the patient-clinician interaction through patient-targeted educational materials or shared decision-making tools are effective in decreasing the use of low-value care. Clinicians and policymakers should consider engaging patients within initiatives that seek to reduce low-value care.
Open Science Framework (https://osf.io/6fsxm).
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