A novel CCM2 variant in a family with non-progressive cognitive complaints and cerebral microbleeds

Lobar cerebral microbleeds are most often sporadic and associated with Alzheimer's disease. The aim of our study was to identify the underlying genetic defect in a family with cognitive complaints and multiple lobar microbleeds and a positive family history for early onset Alzheimer's disease. We performed exome sequencing followed by Sanger sequencing for validation purposes on genomic DNA of three siblings with cognitive complaints, reduced amyloid-beta-42 in CSF and multiple cerebral lobar microbleeds. We checked for the occurrence of the variant in a cohort of 363 patients with early onset dementia and/or microbleeds. A novel frameshift variant (c.236_237delAC) generating a premature stop codon in the CCM2 gene shared by all three siblings was identified. Pathogenicity of the variant was supported by the presence of cerebral cavernous malformations in two of the siblings and by the absence of the variant exome variant databases. Two siblings were homozygous for APOE-ϵ4; one heterozygous. The cognitive complaints, reduced amyloid-beta-42 in CSF and microbleeds suggest preclinical Alzheimer's disease, but the stability of the cognitive complaints does not. We hypothesize that the phenotype in this family may be due to a combination of the CCM2 variant and the APOE status. © 2016 The Authors. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics Published by Wiley Periodicals, Inc

Rare Genetic Variant in SORL1 May Increase Penetrance of Alzheimer's Disease in a Family with Several Generations of APOE-ɛ4 Homozygosity

BACKGROUND: The major genetic risk factor for late onset Alzheimer's disease (AD) is the APOE-ɛ4 allele. However, APOE-ɛ4 homozygosity is not fully penetrant, suggesting co-occurrence of additional genetic variants. OBJECTIVE: To identify genetic factors that, next to APOE-ɛ4 homozygosity, contribute to the development of AD. METHODS: We identified a family with nine AD patients spanning four generations, with an inheritance pattern suggestive of autosomal dominant AD, with no variants in PSEN1, PSEN2, or APP. We collected DNA from four affected and seven unaffected family members and performed exome sequencing on DNA from three affected and one unaffected family members. RESULTS: All affected family members were homozygous for the APOE-ɛ4 allele. Statistical analysis revealed that AD onset in this family was significantly earlier than could be expected based on APOE genotype and gender. Next to APOE-ɛ4 homozygosity, we found that all four affected family members carried a rare variant in the VPS10 domain of the SORL1 gene, associated with AβPP processing and AD risk. Furthermore, three of four affected family members carried a rare variant in the TSHZ3 gene, also associated with AβPP processing. Affected family members presented between 61 and 74 years, with variable presence of microbleeds/cerebral amyloid angiopathy and electroencephalographic abnormalities. CONCLUSION: We hypothesize that next to APOE-ɛ4 homozygosity, impaired SORL1 protein function, and possibly impaired TSHZ3 function, further disturbed Aβ processing. The convergence of these genetic factors over several generations might clarify the increased AD penetrance and the autosomal dominant-like inheritance pattern of AD as observed in this family

Spatiotemporal dynamics of re-innervation and hyperinnervation patterns by uninjured CGRP fibers in the rat foot sole epidermis after nerve injury

The epidermis is innervated by fine nerve endings that are important in mediating nociceptive stimuli. However, their precise role in neuropathic pain is still controversial. Here, we have studied the role of epidermal peptidergic nociceptive fibers that are located adjacent to injured fibers in a rat model of neuropathic pain. Using the Spared Nerve Injury (SNI) model, which involves complete transections of the tibial and common peroneal nerve while sparing the sural and saphenous branches, mechanical hypersensitivity was induced of the uninjured lateral (sural) and medial (saphenous) area of the foot sole. At different time points, a complete foot sole biopsy was taken from the injured paw and processed for Calcitonin Gene-Related Peptide (CGRP) immunohistochemistry. Subsequently, a novel 2D-reconstruction model depicting the density of CGRP fibers was made to evaluate the course of denervation and re-innervation by uninjured CGRP fibers. The results show an increased density of uninjured CGRP-IR epidermal fibers on the lateral and medial side after a SNI procedure at 5 and 10 weeks. Furthermore, although in control animals the density of epidermal CGRP-IR fibers in the footpads was lower compared to the surrounding skin of the foot, 10 weeks after the SNI procedure, the initially denervated footpads displayed a hyper-innervation. These data support the idea that uninjured fibers may play a considerable role in development and maintenance of neuropathic pain and that it is important to take larger biopsies to test the relationship between innervation of injured and uninjured nerve areas

Where Does Mediator Bind In Vivo?

Background: The Mediator complex associates with RNA polymerase (Pol) II, and it is recruited to enhancer regions by activator proteins under appropriate environmental conditions. However, the issue of Mediator association in yeast cells is controversial. Under optimal growth conditions (YPD medium), we were unable to detect Mediator at essentially any S. cerevisiae promoter region, including those supporting very high levels of transcription. In contrast, whole genome microarray experiments in synthetic complete (SC) medium reported that Mediator associates with many genes at both promoter and coding regions. Principal Findings: As assayed by chromatin immunoprecipitation, we show that there are a small number of Mediator targets in SC medium that are not observed in YPD medium. However, most Mediator targets identified in the genome-wide analysis are false positives that arose for several interrelated reasons: the use of overly lenient cut-offs; artifactual differences in apparent IP efficiencies among different genomic regions in the untagged strain; low fold-enrichments making it difficult to distinguish true Mediator targets from false positives that occur in the absence of the tagged Mediator protein. Lastly, apparent Mediator association in highly active coding regions is due to a non-specific effect on accessibility due to the lack of nucleosomes, not to a specific association of Mediator. Conclusions: These results indicate that Mediator does not bind to numerous sites in the yeast genome, but rathe

Stochastic signalling rewires the interaction map of a multiple feedback network during yeast evolution

During evolution, genetic networks are rewired through strengthening or weakening their interactions to develop new regulatory schemes. In the galactose network, the GAL1/GAL3 paralogues and the GAL2 gene enhance their own expression mediated by the Gal4p transcriptional activator. The wiring strength in these feedback loops is set by the number of Gal4p binding sites. Here we show using synthetic circuits that multiplying the binding sites increases the expression of a gene under the direct control of an activator, but this enhancement is not fed back in the circuit. The feedback loops are rather activated by genes that have frequent stochastic bursts and fast RNA decay rates. In this way, rapid adaptation to galactose can be triggered even by weakly expressed genes. Our results indicate that nonlinear stochastic transcriptional responses enable feedback loops to function autonomously, or contrary to what is dictated by the strength of interactions enclosing the circuit

Tumor to normal single-cell mRNA comparisons reveal a pan-neuroblastoma cancer cell

Neuroblastoma is a childhood cancer that resembles developmental stages of the neural crest. It is not established what developmental processes neuroblastoma cancer cells represent. Here, we sought to reveal the phenotype of neuroblastoma cancer cells by comparing cancer (n = 19,723) with normal fetal adrenal single-cell transcriptomes (n = 57,972). Our principal finding was that the neuroblastoma cancer cell resembled fetal sympathoblasts, but no other fetal adrenal cell type. The sympathoblastic state was a universal feature of neuroblastoma cells, transcending cell cluster diversity, individual patients, and clinical phenotypes. We substantiated our findings in 650 neuroblastoma bulk transcriptomes and by integrating canonical features of the neuroblastoma genome with transcriptional signals. Overall, our observations indicate that a pan-neuroblastoma cancer cell state exists, which may be attractive for novel immunotherapeutic and targeted avenues

Adr1 and Cat8 Mediate Coactivator Recruitment and Chromatin Remodeling at Glucose-Regulated Genes

Adr1 and Cat8 co-regulate numerous glucose-repressed genes in S. cerevisiae, presenting a unique opportunity to explore their individual roles in coactivator recruitment, chromatin remodeling, and transcription.We determined the individual contributions of Cat8 and Adr1 on the expression of a cohort of glucose-repressed genes and found three broad categories: genes that need both activators for full derepression, genes that rely mostly on Cat8 and genes that require only Adr1. Through combined expression and recruitment data, along with analysis of chromatin remodeling at two of these genes, ADH2 and FBP1, we clarified how these activators achieve this wide range of co-regulation. We find that Adr1 and Cat8 are not intrinsically different in their abilities to recruit coactivators but rather, promoter context appears to dictate which activator is responsible for recruitment to specific genes. These promoter-specific contributions are also apparent in the chromatin remodeling that accompanies derepression: ADH2 requires both Adr1 and Cat8, whereas, at FBP1, significant remodeling occurs with Cat8 alone. Although over-expression of Adr1 can compensate for loss of Cat8 at many genes in terms of both activation and chromatin remodeling, this over-expression cannot complement all of the cat8Delta phenotypes.Thus, at many of the glucose-repressed genes, Cat8 and Adr1 appear to have interchangeable roles and promoter architecture may dictate the roles of these activators

Single cell derived mRNA signals across human kidney tumors.

Tumor cells may share some patterns of gene expression with their cell of origin, providing clues into the differentiation state and origin of cancer. Here, we study the differentiation state and cellular origin of 1300 childhood and adult kidney tumors. Using single cell mRNA reference maps of normal tissues, we quantify reference "cellular signals" in each tumor. Quantifying global differentiation, we find that childhood tumors exhibit fetal cellular signals, replacing the presumption of "fetalness" with a quantitative measure of immaturity. By contrast, in adult cancers our assessment refutes the suggestion of dedifferentiation towards a fetal state in most cases. We find an intimate connection between developmental mesenchymal populations and childhood renal tumors. We demonstrate the diagnostic potential of our approach with a case study of a cryptic renal tumor. Our findings provide a cellular definition of human renal tumors through an approach that is broadly applicable to human cancer

Med5(Nut1) and Med17(Srb4) Are Direct Targets of Mediator Histone H4 Tail Interactions

The Mediator complex transmits activation signals from DNA bound transcription factors to the core transcription machinery. In addition to its canonical role in transcriptional activation, recent studies have demonstrated that S. cerevisiae Mediator can interact directly with nucleosomes, and their histone tails. Mutations in Mediator subunits have shown that Mediator and certain chromatin structures mutually impact each other structurally and functionally in vivo. We have taken a UV photo cross-linking approach to further delineate the molecular basis of Mediator chromatin interactions and help determine whether the impact of certain Mediator mutants on chromatin is direct. Specifically, by using histone tail peptides substituted with an amino acid analog that is a UV activatible crosslinker, we have identified specific subunits within Mediator that participate in histone tail interactions. Using Mediator purified from mutant yeast strains we have evaluated the impact of these subunits on histone tail binding. This analysis has identified the Med5 subunit of Mediator as a target for histone tail interactions and suggests that the previously observed effect of med5 mutations on telomeric heterochromatin and silencing is direct

The positions of TFIIF and TFIIE in the RNA polymerase II transcription preinitiation complex.

We incorporated the non-natural photoreactive amino acid p-benzoyl-L-phenylalanine (Bpa) into the RNA polymerase II (Pol II) surface surrounding the central cleft formed by the Rpb1 and Rpb2 subunits. Photo-cross-linking of preinitiation complexes (PICs) with these Pol II derivatives and hydroxyl-radical cleavage assays revealed that the TFIIF dimerization domain interacts with the Rpb2 lobe and protrusion domains adjacent to Rpb9, while TFIIE cross-links to the Rpb1 clamp domain on the opposite side of the Pol II central cleft. Mutations in the Rpb2 lobe and protrusion domains alter both Pol II-TFIIF binding and the transcription start site, a phenotype associated with mutations in TFIIF, Rpb9 and TFIIB. Together with previous biochemical and structural studies, these findings illuminate the structural organization of the PIC and the network of protein-protein interactions involved in transcription start site selection