Human monoclonal antibodies to HIV-1 gp140 from mice bearing YAC-based human immunoglobulin transloci.

Mice carrying human immunoglobulin transloci were immunised with HIV-1 gp140 antigen to gain insight into the range and nature of human monoclonal antibodies (mAbs) that can be elicited from such humanised mice. Using five-feature mice that harbour YAC-based germline-configuration human IgM, Igκ and Igλ transloci in a mouse background disrupted for endogenous mouse IgH and Igκ expression, gp140-specific human IgM mAbs were readily elicited following serial immunisation. These mAbs were converted to human IgG1 format and were found to bind diverse epitopes within gp140, exhibiting high functional affinity for the antigen-typically in the nanomolar or sub-nanomolar range.

HS5 of the human beta-globin locus control region: a developmental stage-specific border in erythroid cells.

Elements with insulator/border activity have been characterized most extensively in Drosophila melanogaster. In vertebrates, the first example of such an element was provided by a hypersensitive site of the chicken beta-globin locus, cHS4. It has been proposed that the homologous site in humans, HS5, functions as a border of the human beta-globin locus.

Intergenic transcription and developmental remodeling of chromatin subdomains in the human beta-globin locus.

Gene activation requires chromatin remodeling complexes, which hyperacetylate histones and enable factor access; however, the targeting mechanisms leading to the establishment and maintenance of large, hyperacetylated DNase-sensitive chromatin domains are unknown. Recent work has shown that histone acetyltransferases are associated with RNA-pol II complexes, suggesting that transcription of chromatin plays a role in chromatin modification. Here we show the human beta-globin locus is divided into three differentially activated chromatin subdomains.

Transcriptional activation by hypersensitive site three of the human beta-globin locus control region in murine erythroleukemia cells.

In this paper we describe a complete deletional analysis of hypersensitive site three (HS3) of the human beta-globin Locus Control Region (LCR). The previously defined core fragment consists of 6 footprinted regions, with multiple binding sites for the erythroid-specific factor GATA-1 and G-rich motifs that can interact with ubiquitous factors such as Sp1 and TEF-2. We show in this paper that the 5' half of this fragment is the most important for activity in murine erythroleukemia (MEL) cells.

The chicken beta/epsilon-globin enhancer directs autonomously regulated, high-level expression of the chicken epsilon-globin gene in transgenic mice.

In transiently transfected chicken erythroid cells, beta-like globin gene switching is mediated through differential activation of the cis-linked embryonic epsilon- and adult beta-globin genes by a shared enhancer. Two underlying mechanisms have been proposed: (i) tissue- and stage-specific factors activate the beta-globin promoter in adult erythroid cells (autonomous regulation); and (ii) the epsilon-globin promoter, although transcriptionally competent in both embryonic and adult cells, is suppressed at the adult stage through competition with the beta-globin promoter for interaction with the enhancer (competitive regulation). Analyses of transgenic mice carrying the chicken beta/epsilon-globin locus demonstrated that both genes depended on the enhancer for erythroid expression, but only the epsilon-globin gene exhibited developmentally appropriate transcription at levels comparable to the endogenous mouse globin genes.

Estrogen-inducible and liver-specific expression of the chicken Very Low Density Apolipoprotein II gene locus in transgenic mice.

We have examined the chicken Very Low Density Apolipoprotein II (apoVLDL II) gene locus in transgenic mice. A DNA fragment composed of the transcribed region, 16 kb of 5' flanking and 400 bp of 3' flanking sequences contained all the information sufficient for estrogen-inducible, liver-specific expression of the apoVLDL II gene. The far-upstream region contains a Negative Regulating Element coinciding with a DNaseI-hypersensitive site at -11 kb.

The minimal requirements for activity in transgenic mice of hypersensitive site 3 of the beta globin locus control region.

Proper expression of the human beta-like globin genes is completely dependent on the presence of the locus control region or LCR, a region containing four DNase hypersensitive sites (HS1-4) situated 5' to the structural genes. Linkage of the LCR to a transgene results in copy number-dependent transcription, independent of the site of integration in the host genome. We have analysed a small region of the LCR (HS3) in transgenic animals to determine the minimal interactions that are required for this property.

Each hypersensitive site of the human beta-globin locus control region confers a different developmental pattern of expression on the globin genes.

We have tested the effect of the individual DNase I hypersensitive site (HS) regions of the globin locus control region (LCR) on the developmental expression pattern of the human gamma and beta genes in transgenic mice. The results show that HS3 is the most active site during the embryonic period. It is also the only site capable of high level expression of the gamma genes during fetal hematopoiesis, in a population of cells that are capable of expressing both the gamma and beta genes.

Efficacy and safety of transcutaneous low-impedance cardiac pacing in human volunteers using conventional polymeric defibrillation pads.

Study Objectives: To assess the safety and efficacy of transcutaneous cardiac pacing using low-impedance defibrillation-type, self-adhesive polymer electrode pads positioned in the same anatomic sites typical of such pad placement in emergency defibrillation attempts.

Design: Prospective, randomized, single-blinded normal subject investigation.

Methods: Thirty healthy unmedicated adult volunteers of both sexes were paced transcutaneously to the threshold of capture and beyond by an intensity factor of 125%.

Hypersensitive site 4 of the human beta globin locus control region.

The Locus Control Region (LCR) of the human beta globin gene domain is defined by four erythroid-specific DNasel hypersensitive sites (HSS) located upstream of this multigene cluster. The LCR confers copy number dependent high levels of erythroid specific expression to a linked transgene, independent of the site of integration. To assess the role of the individual hypersensitive sites of the LCR, we have localized HSS4 to a 280bp fragment that is functional both in murine erythroleukaemia (MEL) cells and in transgenic mice.