The rockefeller university

Activation-induced cytidine deaminase (AID) initiates antibody gene diversification by creating U:G mismatches. However, AID is not specific for antibody genes; Off-target lesions can activate oncogenes or cause chromosome translocations. Despite its importance in these transactions little is known about how AID finds its targets. We performed an shRNA screen to identify factors required for class switch recombination (CSR) of antibody loci. We found that Spt5, a factor associated with stalled RNA polymerase II (Pol II) and single stranded DNA (ssDNA), is required for CSR. Spt5 interacts with AID, it facilitates association between AID and Pol II, and AID recruitment to its Ig and non-Ig targets. ChIP-seq experiments reveal that Spt5 colocalizes with AID and stalled Pol II. Further, Spt5 accumulation at sites of Pol II stalling is predictive of AID-induced mutation. We propose that AID is targeted to sites of Pol II stalling in part via its association with Spt5.

Mammalian Rap1, a TRF2-interacting protein in the telomeric shelterin complex, was recently shown to repress homology-directed repair at chromosome ends. In addition, Rap1 plays a role in transcriptional regulation and NF kappa B signaling. Rap1 is unique among the components of shelterin in that it is conserved in budding yeast and has non-telomeric functions. Comparison of mammalian Rap1 to the Rap1 proteins of several budding yeasts and fission yeast reveal both striking similarities and notable differences. The protean nature of Rap1 is best understood by viewing it as an adaptor that can mediate a variety of protein-protein and protein-DNA interactions depending on the organism and the complex in which it is functioning.

Handed asymmetry in organ shape and positioning is a common feature among bilateria, yet little is known about the morphogenetic mechanisms underlying left-right (LR) organogenesis. We utilize the directional 360 degrees clockwise rotation of genitalia in Drosophila to study LA-dependent organ looping. Using time-lapse imaging, we show that rotation of genitalia by 360 degrees results from an additive process involving two ring-shaped domains, each undergoing 180 degrees rotation. Our results show that the direction of rotation for each ring is autonomous and strictly depends on the LR determinant myosin ID (MyoID). Specific inactivation of MyoID in one domain causes rings to rotate in opposite directions and thereby cancels out the overall movement. We further reveal a specific pattern of apoptosis at the ring boundaries and show that local cell death is required for the movement of each domain, acting as a brake-releaser. These data indicate that organ looping can proceed through an incremental mechanism coupling LR determination and apoptosis. Furthermore, they suggest a model for the stepwise evolution of genitalia posture in Diptera, through the emergence and duplication of a 180 degrees LR module.

Accumulation of beta-amyloid (A beta) and loss of synapses are hallmarks of Alzheimer's disease (AD). How synaptic activity relates to A beta accumulation and loss of synapses is a current topic of major interest. Synaptic activation promotes A beta secretion, and chronic reduction of synaptic activity reduced A beta plaques in an AD transgenic mouse model. This suggested beneficial effects of reducing synaptic activity in AD. We now show that reduced synaptic activity causes detrimental effects on synapses and memory despite reducing plaques using two different models of chronic synaptic inhibition: deafferentation of the barrel cortex and administration of benzodiazepine. An interval of prolonged synaptic inhibition exacerbated loss of synaptophysin compared with synaptically more active brain in AD transgenic but not wild-type mice. Furthermore, an interval of benzodiazepine treatment, followed by a washout period, exacerbated memory impairment in AD transgenic mice. Exacerbation of synaptic and behavioral abnormalities occurred in the setting of reduced A beta plaques but elevated intraneuronal A beta immunoreactivity. These data support beneficial effects of synaptic activation on A beta-related synaptic and behavioral impairment in AD.

CLC proteins transport chloride (Cl-) ions across cell membranes to control the electrical potential of muscle cells, transfer electrolytes across epithelia, and control the pH and electrolyte composition of intracellular organelles. Some members of this protein family are Cl- ion channels, whereas others are secondary active transporters that exchange Cl- ions and protons (H+) with a 2: 1 stoichiometry. We have determined the structure of a eukaryotic CLC transporter at 3.5 angstrom resolution. Cytoplasmic cystathionine beta-synthase (CBS) domains are strategically positioned to regulate the ion-transport pathway, and many disease-causing mutations in human CLCs reside on the CBS-transmembrane interface. Comparison with prokaryotic CLC shows that a gating glutamate residue changes conformation and suggests a basis for 2: 1 Cl-/H+ exchange and a simple mechanistic connection between CLC channels and transporters.

A hallmark of mitosis is the appearance of high levels of histone phosphorylation, yet the roles of these modifications remain largely unknown. Here, we demonstrate that histone H3 phosphorylated at threonine 3 is directly recognized by an evolutionarily conserved binding pocket in the BIR domain of Survivin, which is a member of the chromosomal passenger complex (CPC). This binding mediates recruitment of the CPC to chromosomes and the resulting activation of its kinase subunit Aurora B. Consistently, modulation of the kinase activity of Haspin, which phosphorylates H3T3, leads to defects in the Aurora B-dependent processes of spindle assembly and inhibition of nuclear reformation. These findings establish a direct cellular role for mitotic histone H3T3 phosphorylation, which is read and translated by the CPC to ensure accurate cell division.

Objective To test the hypothesis that predisposition to childhood herpes simplex virus (HSV) type 1 encephalitis (HSE) may be determined in part by human genetic factors. Study design A genetic epidemiologic survey of childhood HSE (onset at age 3 months to 15 years) over a 20-year period (1985-2004) was conducted throughout France (comprising 29 university hospital neuropediatric centers). A total of 85 children fulfilled the diagnostic criteria for inclusion. Family and personal histories were obtained by face-to-face interview for 51 patients. Results No familial cases of HSE were identified in our survey; however, a high proportion (20%) of the children interviewed had a relevant family history: parental consanguinity (12% of patients), early-onset herpetic keratitis in a first-degree relative (6%), or both (2%). The narrow window of high susceptibility to HSE before age 3 years (62% of patients) further indicates that predisposition to HSE is tightly age-dependent. Conclusions This survey suggests that childhood HSE, although sporadic, may result from Mendelian predisposition (from autosomal recessive susceptibility in particular), at least in some children. There likely is incomplete penetrance, however, which may reflect, at least in part, the impact of age at the time of HSV-1 infection. (J Pediatr 2010; 157:623-9).

The goal of this study is to examine the expression pattern of orexigenic peptides, orexin (OX) and melanin-concentrating hormone (MCH), in the perifornical lateral hypothalamus (PFLH) in subpopulations of Sprague-Dawley rats differing in their propensity to overconsume a high-fat diet. Immediately after an initial 5-day screening test that predicts long-term consumption, rats identified as high-fat consumers (HFC), ingesting 35% more calories of a high-fat relative to low-fat chow diet, had significantly elevated mRNA expression of OX in the perifornical but not lateral hypothalamic area and of MCH mRNA in both areas, when compared to control rats that consume similar amounts of these diets. This same OX and MCH expression pattern was seen in HFC rats maintained for two weeks on a low-fat chow diet, indicating that increased expression of these orexigenic peptides, occurring independently of the high-fat diet, may be an inherent characteristic of these rats. These HFC rats were also more active and slightly more anxious than controls, as measured by line crossings and time spent in the periphery or middle segments of an open field. Together, these results demonstrate that animals prone to overeating a high-fat diet show a baseline increase in orexigenic peptide expression in the PFLH along with higher behavioral arousal, which together may contribute to their increased consummatory behavior. (C) 2010 Elsevier Inc. All rights reserved.