Differential regulation of sFlt-1 splicing by U2AF65 and JMJD6 in placental-derived and endothelial cells.

Despite years of study, the gestational disorder preeclampsia (PE) remains poorly understood. One proposed mechanism of PE development is increased soluble VEGF receptor-1 (sFlt-1), ultimately causing angiogenic imbalance and endothelial dysfunction. The soluble protein is an alternative splice variant of FLT1, which also encodes for the full-length receptor Flt-1.

Characterization of AN6001, a positive allosteric modulator of α6β2-containing nicotinic acetylcholine receptors.

α6β2-Containing nicotinic acetylcholine receptors (α6β2* nAChRs) are predominantly expressed in midbrain dopaminergic neurons, including substantia nigra pars compacta (SNc) neurons and their projections to striatal regions, where they regulate dopamine release and nigrostriatal activity. It is well established that nAChR agonists exert protection against dopaminergic neurotoxicity in cellular assays and parkinsonian animal models. Historically, drug development in the nAChR field has been mostly focused on development of selective agonists and positive allosteric modulators (PAMs) for the predominant neuronal nAChRs, α7 and α4β2.

PARP1-dependent eviction of the linker histone H1 mediates immediate early gene expression during neuronal activation.

Neuronal stimulation leads to immediate early gene (IEG) expression through calcium-dependent mechanisms. In recent years, considerable attention has been devoted to the transcriptional responses after neuronal stimulation, but relatively little is known about the changes in chromatin dynamics that follow neuronal activation. Here, we use fluorescence recovery after photobleaching, biochemical fractionations, and chromatin immunoprecipitation to show that KCl-induced depolarization in primary cultured cortical neurons causes a rapid release of the linker histone H1 from chromatin, concomitant with IEG expression.

Prosthodontic Rehabilitation for a Patient with Down Syndrome: A Clinical Report.

Patients with Down syndrome can present with a variety of oral manifestations such as hypodontia, periodontal disease, premature tooth loss, reduced salivary flow, crowding of teeth in both arches, and decreased occlusal vertical dimension. The intellectual ability of people with Down syndrome varies widely. They present with a mild-to-moderate intellectual disability that restricts their ability to communicate and adjust to their environment, which can add complexity in the overall dental treatment.

Differential lipid binding of vinculin isoforms promotes quasi-equivalent dimerization.

The main cause of death globally remains debilitating heart conditions, such as dilated cardiomyopathy (DCM) and hypertrophic cardiomyopathy (HCM), which are often due to mutations of specific components of adhesion complexes. Vinculin regulates these complexes and plays essential roles in intercalated discs that are necessary for muscle cell function and coordinated movement and in the development and function of the heart. Humans bearing familial or sporadic mutations in vinculin suffer from chronic, progressively debilitating DCM that ultimately leads to cardiac failure and death, whereas autosomal dominant mutations in vinculin can also provoke HCM, causing acute cardiac failure.

Photobleaching studies reveal that a single amino acid polymorphism is responsible for the differential binding affinities of linker histone subtypes H1.1 and H1.5.

Mammals express six major somatic linker histone subtypes, all of which display dynamic binding to chromatin, characterized by transient binding at a given location followed by rapid translocation to a new site. Using photobleaching techniques, we systematically measured the exchange rate of all six mouse H1 subtypes to determine their relative chromatin-binding affinity. Two subtypes, H1.

Mechanisms and Functions of Vinculin Interactions with Phospholipids at Cell Adhesion Sites.

The cytoskeletal protein vinculin is a major regulator of cell adhesion and attaches to the cell surface by binding to specific phospholipids. Structural, biochemical, and biological studies provided much insight into how vinculin binds to membranes, what components it recognizes, and how lipid binding is regulated. Here we discuss the roles and mechanisms of phospholipids in regulating the structure and function of vinculin and of its muscle-specific metavinculin splice variant.

Molecular dynamics of histone H1.

The H1 or linker histones bind dynamically to chromatin in living cells via a process that involves transient association with the nucleosome near the DNA entry/exit site followed by dissociation, translocation to a new location, and rebinding. The mean residency time of H1 on any given nucleosome is about a minute, which is much shorter than that of most core histones but considerably longer than that of most other chromatin-binding proteins, including transcription factors. Here we review recent advances in understanding the kinetic pathway of H1 binding and how it relates to linker histone structure and function.

A high-throughput screening campaign for detection of ca(2+)-activated k(+) channel activators and inhibitors using a fluorometric imaging plate reader-based tl(+)-influx assay.

The intermediate-conductance Ca(2+)-activated K(+) channel (KCa3.1) has been proposed to play many physiological roles, and modulators of KCa3.1 activity are potentially interesting as new drugs.

Positive allosteric modulation of GABA-A receptors reduces capsaicin-induced primary and secondary hypersensitivity in rats.

GABA-A receptor positive allosteric modulators (PAMs) mediate robust analgesia in animal models of pathological pain, in part via enhancing injury-induced loss of GABA-A-α2 and -α3 receptor function within the spinal cord. As yet, a lack of clinically suitable tool compounds has prevented this concept being tested in humans. Prior to assessing the efficacy of GABA-A receptor PAMs in a human volunteer pain model we have compared compounds capable of variously modulating GABA-A receptor function in comparable rat models of capsaicin-induced acute nocifensive flinching behaviour and secondary mechanical hypersensitivity.

N- and C-terminal domains determine differential nucleosomal binding geometry and affinity of linker histone isotypes H1(0) and H1c.

Eukaryotic linker or H1 histones modulate DNA compaction and gene expression in vivo. In mammals, these proteins exist as multiple isotypes with distinct properties, suggesting a functional significance to the heterogeneity. Linker histones typically have a tripartite structure composed of a conserved central globular domain flanked by a highly variable short N-terminal domain and a longer highly basic C-terminal domain.

Potential adverse reactions to herbal medicines in patients attending a nephrology clinic in Abu Dhabi, United Arab Emirates.

Objective: To provide data on herbal medicine (HM) use and safety in patients attending a nephrology clinic at Sheikh Khalifa Medical City (SKMC), Abu Dhabi, United Arab Emirates (UAE).

Methods: A prospective, 3-month study between June and September 2007, investigated all patients presenting to the Nephrology Clinic of the Sheikh Khalifa Medical center (SKMC) in Abu Dhabi, UAE. A structured questionnaire determined previous and current HM use, and descriptions of associated adverse reactions.

Nucleosome interaction surface of linker histone H1c is distinct from that of H1(0).

The fully organized structure of the eukaryotic nucleosome remains unsolved, in part due to limited information regarding the binding site of the H1 or linker histone. The central globular domain of H1 is believed to interact with the nucleosome core at or near the dyad and to bind at least two strands of DNA. We utilized site-directed mutagenesis and in vivo photobleaching to identify residues that contribute to the binding of the globular domain of the somatic H1 subtype H1c to the nucleosome.

Prothymosin alpha is a component of a linker histone chaperone.

Linker histone H1 binds with high affinity to naked and nucleosomal DNA in vitro but is rapidly exchanged between chromatin sites in vivo suggesting the involvement of one or more linker histone chaperones. Using permeabilized cells, we demonstrate that the small acidic protein prothymosin alpha (ProTalpha) can facilitate H1 displacement from and deposition onto the native chromatin template. Depletion of ProTalpha levels in vivo by siRNA-mediated mRNA degradation resulted in a decreased rate of exchange of linker histones as assayed by photobleaching techniques.

Dissecting the binding mechanism of the linker histone in live cells: an integrated FRAP analysis.

The linker histone H1 has a fundamental role in DNA compaction. Although models for H1 binding generally involve the H1 C-terminal tail and sites S1 and S2 within the H1 globular domain, there is debate about the importance of these binding regions and almost nothing is known about how they work together. Using a novel fluorescence recovery after photobleaching (FRAP) procedure, we have measured the affinities of these regions individually, in pairs, and in the full molecule to demonstrate for the first time that binding among several combinations is cooperative in live cells.

Global modulation of chromatin dynamics mediated by dephosphorylation of linker histone H1 is necessary for erythroid differentiation.

Differentiation of metazoan cells involves dramatic changes in gene expression patterns and proliferative capacity driven primarily by epigenetic mechanisms. Here we used in vivo photobleaching techniques and biochemical assays to investigate the contribution of alterations in chromatin dynamics to the differentiation of murine erythroleukemia (MEL) cells, a model system for erythroid development. As MEL cells differentiate the binding affinity of all linker histone variants increases, indicative of an overall decrease in chromatin flexibility.

Mapping the interaction surface of linker histone H1(0) with the nucleosome of native chromatin in vivo.

H1 linker histones stabilize the nucleosome, limit nucleosome mobility and facilitate the condensation of metazoan chromatin. Here, we have combined systematic mutagenesis, measurement of in vivo binding by photobleaching microscopy, and structural modeling to determine the binding geometry of the globular domain of the H1(0) linker histone variant within the nucleosome in unperturbed, native chromatin in vivo. We demonstrate the existence of two distinct DNA-binding sites within the globular domain that are formed by spatial clustering of multiple residues.

Hyperdynamic plasticity of chromatin proteins in pluripotent embryonic stem cells.

Differentiation of embryonic stem (ES) cells from a pluripotent to a committed state involves global changes in genome expression patterns. Gene activity is critically determined by chromatin structure and interactions of chromatin binding proteins. Here, we show that major architectural chromatin proteins are hyperdynamic and bind loosely to chromatin in ES cells.

Differential in vivo binding dynamics of somatic and oocyte-specific linker histones in oocytes and during ES cell nuclear transfer.

The embryonic genome is formed by fusion of a maternal and a paternal genome. To accommodate the resulting diploid genome in the fertilized oocyte dramatic global genome reorganizations must occur. The higher order structure of chromatin in vivo is critically dependent on architectural chromatin proteins, with the family of linker histone proteins among the most critical structural determinants.

Global nature of dynamic protein-chromatin interactions in vivo: three-dimensional genome scanning and dynamic interaction networks of chromatin proteins.

Genome structure and gene expression depend on a multitude of chromatin-binding proteins. The binding properties of these proteins to native chromatin in intact cells are largely unknown. Here, we describe an approach based on combined in vivo photobleaching microscopy and kinetic modeling to analyze globally the dynamics of binding of chromatin-associated proteins in living cells.

DNA intercalators differentially affect chromatin structure and DNA replication in Xenopus egg extract.

In this paper, we describe a scheme utilizing the Xenopus egg extract system to simultaneously evaluate DNA-interacting drugs as potential anti-cancer agents and gain insights into the mechanisms of drug action. We studied two DNA intercalators, daunomycin (DM), a cancer chemotherapeutic, and ethidium bromide (EtBr), a compound with no reported therapeutic value. Consistent with our earlier report, we find that DM inhibits DNA replication in a concentration-dependent manner.

Evaluation of community pharmacists' recommendations to standardized patient scenarios.

Background: Recent trends in the global nonprescription drug market show a massive increase in medicine availability to the general public. This places greater responsibility on community pharmacists to ensure that patients exercise self-care appropriately. This small study examined pharmacist performance in facilitating patient self-care.