Thymocyte-specific truncation of the deubiquitinating domain of CYLD impairs positive selection in a NF-kappaB essential modulator-dependent manner

The cylindromatosis tumor suppressor gene (Cyld) encodes a deubiquitinating enzyme (CYLD) with immunoregulatory function. In this study, we evaluated the role of Cyld in T cell ontogeny by generating a mouse (Cyld(Delta9)) with a thymocyte-restricted Cyld mutation that causes a C-terminal truncation of the protein and reciprocates catalytically inactive human mutations. Mutant mice had dramatically reduced single positive thymocytes and a substantial loss of peripheral T cells. The analyses of polyclonal and TCR-restricted thymocyte populations possessing the mutation revealed a significant block in positive selection and an increased occurrence of apoptosis at the double-positive stage. Interestingly, in the context of MHC class I and II restricted TCR transgenes, lack of functional CYLD caused massive deletion of thymocytes that would have been positively selected, which is consistent with an impairment of positive selection. Biochemical analysis revealed that Cyld(Delta9) thymocytes exhibit abnormally elevated basal activity of NF-kappaB and JNK. Most importantly, inactivation of NF-kappaB essential modulator fully restored the NF-kappaB activity of Cyld(Delta9) thymocytes to physiologic levels and rescued their developmental and survival defect. This study identifies a fundamental role for functional CYLD in establishing the proper threshold of activation for thymocyte selection by a mechanism dependent on NF-kappaB essential modulator

Talin1 dysfunction is genetically linked to systemic capillary leak syndrome.

Systemic capillary leak syndrome (SCLS) is a rare life-threatening disorder due to profound vascular leak. The trigger and the cause of the disease is currently unknown and there is no specific treatment. Here, we identified a rare heterozygous splice-site variant in the TLN1 gene in a familial SCLS case, suggestive of autosomal dominant inheritance with incomplete penetrance. Talin1 has a key role in cell adhesions by activating and linking integrins to the actin cytoskeleton. This variant causes in-frame skipping of exon 54 and is predicted to affect talin’s c-terminal actin binding site (ABS3). Modelling the SCLS-TLN1 variant by mimicking the actin-binding disruption in TLN1 heterozygous endothelial cells resulted in disorganized endothelial adherens junctions. Mechanistically, we established that disruption of talin’s ABS3 sequestrates talin’s interacting partner, vinculin, at cell-extracellular matrix adhesions, leading to destabilization of the endothelial barrier. We propose that pathogenic variant in TLN1 underlie SCLS, providing insight into the molecular mechanism of the disease which can be explored for future therapeutic interventions

Evidence for alternative splicing and developmental regulation of the Drosophila melanogaster Mgat2 (N-acetylglucosaminyltransferase II) gene

We present a molecular study of the Drosophila melanogaster Mgat2 gene that codes for the N-acetylglucosaminyliransferase II. Isolation and analysis of two cDNA clones indicated that the gene is alternatively spliced, generating two transcripts of 3.3 and 2.7 kb. Developmental specificity was observed between the two alternative transcripts during the major developmental stages of D. melanogaster. The deduced amino acid sequences of the two proteins show significant homology to the equivalent mammalian proteins, especially in the carboxyterminal region. In situ hybridizations in embryos and embryonic imaginal discs showed that the gene is expressed mainly but not exclusively in the brain. Published by Elsevier Inc

Suppression of tau‐induced phenotypes by vitamin E demonstrates the dissociation of oxidative stress and phosphorylation in mechanisms of tau toxicity

Various lines of evidence implicate oxidative stress in the pathogenic mechanism(s) underpinning tauopathies. Consequently, antioxidant therapies have been considered in clinical practice for the treatment of tauopathies such as Alzheimer's disease (AD), but with mixed results. We and others have previously reported increased protein oxidation upon expression of both human 0N3R (hTau 0N3R) and 0N4R (hTau 0N4R) tau in vivo. Building on these studies, we demonstrate here the suppression of hTau 0N3R associated phenotypes in Drosophila melanogaster after treatment with vitamin C or vitamin E. Curiously the rescue of phenotype was seen without alteration in total tau level or alteration in phosphorylation at a number of disease-associated sites. Moreover, treatment with paraquat, a pro-oxidant drug, did not exacerbate the hTau 0N3R phenotypes. This result following paraquat treatment is reminiscent of our previous findings with hTau 0N4R which also causes greater oxidative stress when compared to hTau 0N3R but has a milder phenotype. Collectively our data imply that the role of oxidative stress in tau-mediated toxicity is not straight forward and there may be isoform-specific effects as well as contribution of other factors. This may explain the ambiguous effects of anti-oxidant treatments on clinical outcome in dementia patients. (Figure presented.). </p

The transcription factor BCL-6 controls early development of innate-like T cells

Innate T cells, including invariant natural killer T (iNKT) and mucosal-associated innate T (MAIT) cells, are a heterogeneous T lymphocyte population with effector properties preprogrammed during their thymic differentiation. How this program is initiated is currently unclear. Here, we show that the transcription factor BCL-6 was transiently expressed in iNKT cells upon exit from positive selection and was required for their proper development beyond stage 0. Notably, development of MAIT cells was also impaired in the absence of Bcl6. BCL-6-deficient iNKT cells had reduced expression of genes that were associated with the innate T cell lineage, including Zbtb16, which encodes PLZF, and PLZF-targeted genes. BCL-6 contributed to a chromatin accessibility landscape that was permissive for the expression of development-related genes and inhibitory for genes associated with naive T cell programs. Our results revealed new functions for BCL-6 and illuminated how this transcription factor controls early iNKT cell development. © 2020, The Author(s), under exclusive licence to Springer Nature America, Inc

Repression of differentiation genes by Hes transcription factors fuels neural tumour growth in Drosophila

Background: Neural stem cells (NSC) in divide asymmetrically to generate one cell that retains stem cell identity and another that is routed to differentiation. Prolonged mitotic activity of the NSCs gives rise to the plethora of neurons and glial cells that wire the brain and nerve cord. Genetic insults, such as excess of Notch signaling, perturb the normal NSC proliferation programs and trigger the formation of NSC hyperplasias, which can subsequently progress to malignancies. Hes proteins are crucial mediators of Notch signaling, and in the NSC context they act by repressing a cohort of early pro-differentiation transcription factors. Downregulation of these pro-differentiation factors makes NSC progeny cells susceptible to adopting an aberrant stem cell program. We have recently shown that Hes overexpression in Drosophila leads to NSC hyperplasias that progress to malignant tumours after allografting to adult hosts. Methods: We have combined genetic analysis, tissue allografting and transcriptomic approaches to address the role of Hes genes in NSC malignant transformation. Results: We show that the E (spl) genes are important mediators in the progression of Notch hyperplasias to malignancy, since allografts lacking the E (spl) genes grow much more slowly. We further present RNA profiling of Hes-induced tumours at two different stages after allografting. We find that the same cohort of differentiation-promoting transcription factors that are repressed in the primary hyperplasias continue to be downregulated after transplantation. This is accompanied by an upregulation of stress-response genes and metabolic reprogramming. Conclusions: The combination of dedifferentiation and cell physiology changes most likely drive tumour growth