Biomedical subjects

M Artinger

Publications and source records attributed to M Artinger.

6 recordsLinked to original sources

PubMed2001

Hierarchical cleavage of focal adhesion kinase by caspases alters signal transduction during apoptosis of intestinal epithelial cells.

BACKGROUND & AIMS: Purified intestinal epithelial cells die of detachment-induced apoptosis due to loss of cell anchorage during isolation. Anchorage-dependent cells form focal adhesions, sites of enhanced cell-matrix attachment that confer survival signals. Focal adhesion kinase (FAK), a component of the focal adhesion signaling complex, transduces these antiapoptotic signals. In this report, the molecular events leading to cleavage of FAK by caspases during apoptosis and its functional implications are defined. METHODS: Cytosolic extracts of human intestinal epithelial cells undergoing detachment-induced apoptosis were analyzed by Western blotting, immunoprecipitation, and kinase assay. RESULTS: FAK is cleaved by the ordered proteolytic activity of 2 different members of the caspase-3 family. The first cleavage is mediated by caspase-3, generating a 94/92-kilodalton-terminal fragment, which is processed by caspase-6 to an 84-kilodalton fragment. After apoptosis is initiated, the level of FAK phosphorylation is rapidly decreased, and the phosphorylation pattern of FAK-associated proteins is dramatically modified, showing significant yet divergent changes in signal transduction. CONCLUSIONS: Cleavage of FAK during apoptosis of normal human cells is an example of the sequential, highly regulated, and coordinate action of caspases that not only dismantle a cell by proteolysis, but also alter the cell's signaling machinery.

Amino Acid Chloromethyl Ketones↗

PubMed1998

Antimorphic goosecoids.

goosecoid (gsc) is a homeobox gene expressed in the Spemann organizer that has been implicated in vertebrate axis formation. Here antimorphic gscs are described. One antimorphic gsc (MTgsc) was fortuitously created by adding 5 myc epitopes to the N terminus of gsc. The other antimorph (VP16gsc) contains the transcriptional activation domain of VP16. mRNA injection of either antimorph inhibits dorsal gastrulation movements and leads to embryos with severe axial defects. They upregulate ventral gene expression in the dorsal marginal zone and inhibit dorsal mesoderm differentiation. Like the VP16 domain, the N-terminal myc tags act by converting wild-type gsc from a transcriptional repressor into an activator. However, unlike MTgsc, VP16gsc is able at low dose to uncouple head from trunk formation, indicating that different antimorphs may elicit distinct phenotypes. The experiments reveal that gsc and/or gsc-related genes function in axis formation and gastrulation. Moreover, this work warns against using myc tags indiscriminately for labeling DNA-binding proteins.

Animals↗

PubMed1998

High throughput analysis of differential gene expression.

Elucidation of the changes in gene expression associated with biological processes is a central problem in biology. Advances in molecular and computational biology have led to the development of powerful, high-throughput methods for the analysis of differential gene expression. These tools have opened up new opportunities in disciplines ranging from cell and developmental biology to drug development and pharmacogenomics. In this review, the attributes of five commonly used differential gene expression methods are discussed: expressed sequence tag (EST) sequencing, cDNA microarray hybridization, subtractive cloning, differential display, and serial analysis of gene expression (SAGE). The application of EST sequencing and microarray hybridization is illustrated by the discovery of novel genes associated with osteoblast differentiation. The application of subtractive cloning is presented as a tool to identify genes regulated in vivo by the transcription factor pax-6. These and other examples illustrate the power of genomics for discovering novel genes that are important in biology and which also represent new targets for drug development. The central theme of the review is that each of the approaches to identifying differentially expressed genes is useful, and that the experimental context and subsequent evaluation of differentially expressed genes are the critical features that determine success.

Animals↗

PubMed1997

Interaction of goosecoid and brachyury in Xenopus mesoderm patterning.

Detailed in situ analyses reveal overlapping expression of gsc and Xbra in the early Spemann's organizer. Coexpression is lost during gastrulation suggesting an interaction between these genes. Ectopic expression of gsc ventrally suppresses endogenous Xbra expression and transcription from Xbra promoter reporter gene constructs. Suppression is mediated, at least partially, by a gsc-binding site within the first 349 bp of the promoter. Xbra reporter gene transcription is also suppressed in the region of endogenous gsc expression, whereas high-level ectopic Xbra expression has no effect on endogenous gsc expression. We suggest that early patterning of the vertebrate mesoderm, like early patterning of the Drosophila embryo, occurs by first establishing broad domains of gene expression which are subsequently refined by intergenic interactions to further delimit tissue boundaries.

Animals↗

PubMed1992

Developmentally regulated expression of alpha 6 integrin in avian embryos.

The distribution pattern of the avian alpha 6 integrin subunit was examined during early stages of development. The results show that this subunit is prevalent in cells of the developing nervous system and muscle. alpha 6 is first observed on neuroepithelial cells of the cranial neural plate and trunk neural tube. With time, immunoreactivity becomes prominent near the lumen and ventrolateral portions of the neural tube, co-distributing with neurons and axons, particularly notable on commissural neurons. The alpha 6 expression pattern is dynamic in the neural tube, with immunoreactivity peaking by embryonic day 6 (stage 30) and decreasing thereafter. The ventral roots and retina exhibit high levels of immunoreactivity throughout development. In the peripheral nervous system, alpha 6 immunoreactivity first appears on a subpopulation of sympathoadrenal cells around the dorsal aorta and later in the dorsal root ganglia shortly after gangliogenesis. Immunoreactivity appears on prospective myotomal cells as the somites delaminate into the dermomyotome and sclerotome, remaining prominent on myoblasts and differentiated muscle at all stages. The mesonephros also has intense immunoreactivity. In the periphery, alpha 6 immunoreactive regions often in proximity to laminin, which is thought to be the ligand of alpha 6 beta 1 integrin.

Animals↗

PubMed1990

Distribution and biochemical characterization of the INO antigen during chick neural crest cell migration.

The INO (inhibitor of neurite outgrowth) antibody recognizes a laminin-heparan sulfate proteoglycan complex and was isolated for its ability to functionally inhibit axonal outgrowth of peripheral neurons. Here, we examine the distribution and biochemical characteristics of INO in the early chick embryo. Because the INO antigen is sensitive to most classical fixation procedures and fixation leads to abundant nuclear staining, the antibody was directly injected into 1.5-2.5-day-old embryos prior to fixation. The distribution of the injected antibody was then observed in cryostat sections by indirect immunofluorescence. Particular attention was focussed upon regions of ongoing neural crest cell migration. The INO antigen was observed along both cranial and trunk neural crest cell migratory pathways. The antigen was seen around the basement membrane surrounding the neural tube and notochord, and underneath the ectoderm and endoderm. In addition, fibrillar staining was observed in the cranial mesenchyme and in both rostral and caudal halves of the somitic sclerotome in the trunk. The distribution pattern was identical to that previously observed for laminin or heparan sulfate proteoglycan. To confirm the nature of the INO antigen, we performed immunoprecipitations of chick embryos ranging from 1.5 to 9 days of incubation. Half of each sample was digested with heparinase prior to SDS-PAGE and silver staining. In material from young embryos, bands of 200 and 180 kD (probably corresponding to the B-chains of laminin) plus two broad smears of bands at 180-150 kD and 130-85 kD were observed without heparinase digestion. Following enzymatic digestion, the 200-kD and 180-kD bands remained, while the smears disappeared and were replaced by numerous low-molecular-weight bands. In contrast to preparations from young embryos, samples taken from embryos at day 3 or beyond did not enter the 8% gel without heparinase digestion, though the banding pattern appeared identical to younger samples after heparinase digestion in the presence or absence of Ca2+. This change in the INO antigen with age could result from an increase in the heparin-side-chains attached to similar core proteins, or from an increase in the stability of the laminin-heparan sulfate proteoglycan containing complex with time.

Animals↗