The rockefeller university

Expansion of a hexanucleotide repeat (HRE), GGGGCC, in the C9ORF72 gene is recognized as the most common cause of familial amyotrophic lateral sclerosis (FALS), frontotemporal dementia (FTD) and ALS-FTD, as well as 5-10% of sporadic ALS. Despite the location of the HRE in the non-coding region (with respect to the main C9ORF72 gene product), dipeptide repeat proteins (DPRs) that are thought to be toxic are translated from the HRE in all three reading frames from both the sense and antisense transcript. Here, we identified a CUG that has a good Kozak consensus sequence as the translation initiation codon. Mutation of this CTG significantly suppressed polyglycine-alanine (GA) translation. GA was translated when the G(4)C(2) construct was placed as the second cistron in a bicistronic construct. CRISPR/Cas9-induced knockout of a non-canonical translation initiation factor, eIF2A, impaired GA translation. Transfection of G(4)C(2) constructs induced an integrated stress response (ISR), while triggering the ISR led to a continuation of translation of GA with a decline in conventional cap-dependent translation. These in vitro observations were confirmed in chick embryo neural cells. The findings suggest that DPRs translated from an HRE in C9ORF72 aggregate and lead to an ISR that then leads to continuing DPR production and aggregation, thereby creating a continuing pathogenic cycle.

Research question: What level of IVF pregnancy success is currently possible in women of extremely advanced age? Design: This study reports on outcomes in women aged 43-51 years at the Centre for Human Reproduction, an academically affiliated private clinical fertility and research centre in New York City. Results: During the study years of 2014-2016, 16 pregnancies were established, all through day 3 transfers. Based on 'intent to treat' (cycle start). clinical pregnancy rates were 4/190 (2.1%), 5/234 (2.1%) and 7/304 (2.3%) and live birth rates were 2/190 (1.1%), 1/234 (0.43%) and 4/304 (1.3%) in 2014, 2015 and 2016, respectively. With reference to embryo transfer, clinical pregnancy rates were 4/140 (2.9%), 5/159 (3.1%) and 7/167 (4.2%) and live birth rates were 2/140 (1.4%), 1/159 (0.63%) and 4/167 (2.4%) for the same years. The results for 2016 also included what are probably the two oldest autologous IVF pregnancies ever reported in the literature. These results were obtained with patient ages, percentage of cycle cancellations and other adverse outcome parameters steadily increasing year by year. Conclusions: Female age above 42 is widely viewed as the ultimate barrier to conception with IVF. Data reported here, although small and preliminary, demonstrate that potential outcomes are better than widely perceived, while pregnancy and live birth rates remain significantly inferior to donor egg recipient cycles. However, for selected women at very advanced ages, especially with higher egg/embryo numbers, autologous oocyte IVF offers a better option than widely acknowledged, if they are given individualized age-specific care. (C) 2018 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

The pathogenesis of degenerative disc disease is a complex and multifactorial process in which genetics, mechanical trauma, altered loading and nutrition present significant etiological factors. Infection of the intervertebral disc with the anaerobic bacterium Propionibacteriumacnes is now also emerging as a potentially new etiological factor. This human commensal bacterium is well known for its long association with the inflammatory skin condition acne vulgaris. A key component of inflammatory responses to P. acnes in acne appears to be interleukin (IL)-1 beta. Similarly, in degenerative disc disease (DDD) there is compelling evidence for the fundamental roles of IL-1 beta in its pathology. We therefore propose that P. acnes involvement in DDD is biologically very plausible, and that IL-1 beta is the key inflammatory mechanism driving the host response to P. acnes infection. Since there is a solid theoretical basis for this phenomenon, we further propose that the relationship between P. acnes infection and DDD is causal.

Tissue homeostasis depends on the ability of tissue-specific adult stem cells to maintain a balance between proliferation and differentiation, as well as ensure DNA damage repair. Here, we use the Drosophila male germline stem cell system to study how a chromatin factor, enhancer of polycomb [E(Pc)], regulates the proliferation-to-differentiation (mitosis-to-meiosis) transition and DNA damage repair. We identified two critical targets of E(Pc). First, E(Pc) represses CycB transcription, likely through modulating H4 acetylation. Second, E(Pc) is required for accumulation of an important germline differentiation factor, Bag-of-marbles (Bam), through post-transcriptional regulation. When E(Pc) is downregulated, increased CycB and decreased Bam are both responsible for defective mitosis-to-meiosis transition in the germline. Moreover, DNA double-strand breaks (DSBs) accumulate upon germline inactivation of E(Pc) under both physiological condition and recovery from heat shock-induced endonuclease expression. Failure of robust DSB repair likely leads to germ cell loss. Finally, compromising the activity of Tip60, a histone acetyltransferase, leads to germline defects similar to E(Pc) loss-of-function, suggesting that E(Pc) acts cooperatively with Tip60. Together, our data demonstrate that E(Pc) has pleiotropic roles in maintaining male germline activity and genome integrity. Our findings will help elucidate the in vivo molecular mechanisms of E(Pc).

beta-Arrestins (beta-arrestin-1 and -2) are multifunctional proteins that play important roles in the regulation of inflammation and cell survival that need to be tightly controlled; however, the mechanism that underlies their gene expression is largely unclear. Here, we demonstrate that -arrestin-1 is a transcriptional target of NF-B. mRNA and protein levels of -arrestin-1 were up-regulated by NF-B inducers. Inhibition of NF-B prevented the up-regulation of -arrestin-1 mRNA, whereas activation of NF-B led to increased -arrestin-1 expression. -Arrestin-1 promoter activity was consistently enhanced upon NF-B activation as a result of the presence of a highly conserved B site. -Arrestin-1, in turn, suppressed the transcriptional activity of NF-B by interfering with the interaction between p65 and p50. -Arrestin-1-deficient mice displayed reduced TNF--induced cell death and increased expression of antiapoptotic genes. Reintroduction of -arrestin-1, but not its mutant, which is unable to interfere with the p65-p50 interaction, into -arrestin-deficient mouse embryonic fibroblasts partially restored sensitivity to TNF--induced cell death. These findings reveal NF-B and -arrestin-1 to be key components of a negative feedback circuit that is necessary to regulate cell death.Li, J., Guo, A., Wang, Q., Li, Y., Zhao, J., Lu, J., Pei, G. NF-B directly regulates beta-arrestin-1 expression and forms a negative feedback circuit in TNF-alpha-induced cell death.

HIV-1 accesses the nuclear DNA of interphase cells via a poorly defined process involving functional interactions between the capsid protein (CA) and nucleoporins (Nups). Here, we show that HIV-1 CA can bind multiple Nups, and that both natural and manipulated variation in Nup levels impacts HIV-1 infection in a manner that is strikingly dependent on cell-type, cell-cycle, and cyclophilin A (CypA). We also show that Nups mediate the function of the antiviral protein MX2, and that MX2 can variably inhibit non-viral NLS function. Remarkably, both enhancing and inhibiting effects of cyclophilin A and MX2 on various HIV-1 CA mutants could be induced or abolished by manipulating levels of the Nup93 subcomplex, the Nup62 subcomplex, NUP88, NUP214, RANBP2, or NUP153. Our findings suggest that several Nup-dependent 'pathways' are variably exploited by HIV-1 to target host DNA in a cell-type, cell-cycle, CypA and CA-sequence dependent manner, and are differentially inhibited by MX2.

The diffusivity of macromolecules in the cytoplasm of eukaryotic cells varies over orders of magnitude and dictates the kinetics of cellular processes. However, a general description that associates the Brownian or anomalous nature of intracellular diffusion to the architectural and biochemical properties of the cytoplasm has not been achieved. Here we measure the mobility of individual fluorescent nanoparticles in living mammalian cells to obtain a comprehensive analysis of cytoplasmic diffusion. We identify a correlation between tracer size, its biochemical nature and its mobility. Inert particles with size equal or below 50 nm behave as Brownian particles diffusing in a medium of low viscosity with negligible effects of molecular crowding. Increasing the strength of non-specific interactions of the nanoparticles within the cytoplasm gradually reduces their mobility and leads to subdiffusive behaviour. These experimental observations and the transition from Brownian to subdiffusive motion can be captured in a minimal phenomenological model.

Biallelic loss-of-function (LOF) mutations of the NCF4 gene, encoding the p40(phox) subunit of the phagocyte NADPH oxidase, have been described in only 1 patient. We report on 24 p40(phox)-deficient patients from 12 additional families in 8 countries. These patients display 8 different in-frame or out-of-frame mutations of NCF4 that are homozygous in 11 of the families and compound heterozygous in another. When overexpressed in NB4 neutrophil-like cells and EBV-transformed B cells in vitro, the mutant alleles were found to be LOF, with the exception of the p.R58C and c.120_134del alleles, which were hypomorphic. Particle-induced NADPH oxidase activity was severely impaired in the patients' neutrophils, whereas PMAinduced dihydrorhodamine-1,2,3 (DHR) oxidation, which is widely used as a diagnostic test for chronic granulomatous disease (CGD), was normal or mildly impaired in the patients. Moreover, the NADPH oxidase activity of EBV-transformed B cells was also severely impaired, whereas that of mononuclear phagocytes was normal. Finally, the killing of Candida albicans and Aspergillus fumigatus hyphae by neutrophils was conserved in these patients, unlike in patients with CGD. The patients suffer from hyperinflammation and peripheral infections, but they do not have any of the invasive bacterial or fungal infections seen in CGD. Inherited p40(phox) deficiency underlies a distinctive condition, resembling a mild, atypical form of CGD.

An essential step in the development of effective antiviral humoral responses is cytokine-triggered class switch recombination resulting in the production of antibodies of a specific isotype. Most viral and parasitic infections in mice induce predominantly IgG2a-specific antibody responses that are stimulated by interferon gamma (IFN-gamma). However, in some mice deficient in IFN-gamma, class switching to IgG2a antibodies is relatively unaffected, indicating that another signal(s) can be generated upon viral or parasitic infections that trigger this response. Here, we found that a single recessive locus, provisionally called IFN-gamma-independent IgG2a (Igii), confers the ability to produce IFN-gamma-independent production of IgG2a antibodies upon retroviral infection. The Igii locus was mapped to chromosome 9 and was found to function in the radiation-resistant compartment. Thus, our data implicate nonhematopoietic cells in activation of antiviral antibody responses in the absence of IFN-gamma. IMPORTANCE Understanding the signals that stimulate antibody production and class switch recombination to specific antibody isotypes is crucial for the development of novel vaccines and adjuvants. While an interferon gamma-mediated switch to the IgG2a isotype upon viral infection in mice has been well established, this investigation reveals a noncanonical, interferon gamma-independent pathway for anti-retroviral antibody production and IgG2a class switch recombination that is controlled by a single recessive locus. Furthermore, this study indicates that the radiation-resistant compartment can direct antiviral antibody responses, suggesting that detection of infection by nonhematopoietic cells is involved is stimulating adaptive immunity.

Background: Division of labor between reproductive queens and workers that perform brood care is a hallmark of insect societies. However, studies of the molecular basis of this fundamental dichotomy are limited by the fact that the caste of an individual cannot typically be experimentally manipulated at the adult stage. Here we take advantage of the unique biology of the clonal raider ant, Ooceraea biroi, to study brain gene expression dynamics during experimentally induced transitions between reproductive and brood care behavior. Results: Introducing larvae that inhibit reproduction and induce brood care behavior causes much faster changes in adult gene expression than removing larvae. In addition, the general patterns of gene expression differ depending on whether ants transition from reproduction to brood care or vice versa, indicating that gene expression changes between phases are cyclic rather than pendular. Finally, we identify genes that could play upstream roles in regulating reproduction and behavior because they show large and early expression changes in one or both transitions. Conclusions: Our analyses reveal that the nature and timing of gene expression changes differ substantially depending on the direction of the transition, and identify a suite of promising candidate molecular regulators of reproductive division of labor that can now be characterized further in both social and solitary animal models. This study contributes to understanding the molecular regulation of reproduction and behavior, as well as the organization and evolution of insect societies.