The juxtamembrane regions of the epidermal growth factor receptor and gp185erbB-2 determine the specificity of signal transduction.

The epidermal growth factor receptor (EGFR) and gp185erbB-2 are closely related tyrosine kinases. Despite extensive sequence and structural homology, these two receptors display quantitative and qualitative differences in their ability to couple with mitogenic signalling pathways. By using chimeric molecules between EGFR and erbB-2, we found that the determinants responsible for the specificity of mitogenic signal transduction are located in the amino-terminal half of the tyrosine kinase domain of either receptor. In the EGFR, mutational analysis within this subdomain revealed that deletion of residues 660 to 667 impaired receptor mitogenic activity without affecting its tyrosine kinase properties. This sequence is therefore likely to contribute to the specificity of substrate recognition by the EGFR kinase.     

Characterization of the receptor for keratinocyte growth factor. Evidence for multiple fibroblast growth factor receptors

Keratinocyte growth factor (KGF) is a member of the fibroblast growth factor (FGF) family. KGF exhibits potent mitogenic activity for a variety of epithelial cell types but is distinct from other known FGFs in that it is not mitogenic for fibroblasts or endothelial cells. We report saturable specific binding of 125I-KGF to surface receptors on intact Balb/MK mouse epidermal keratinocytes. 125I-KGF binding was completed efficiently by acidic FGF (aFGF) but with 20-fold lower efficiency by basic FGF (bFGF). The pattern of 125I-acidic FGF binding and competition on Balb/MK keratinocytes and NIH/3T3 fibroblasts suggests that these cell types possess related but distinct FGF receptors. Scatchard analysis of 125I-KGF binding suggested major and minor high affinity receptor components (KD = 400 and 25 pM, respectively) as well as a third high capacity/low affinity heparin-like component. Covalent affinity cross-linking of 125I-KGF to its receptor on Balb/MK cells revealed two species of 115 and 140 kDa. KGF also stimulated the rapid tyrosine phosphorylation of a 90-kDa protein in Balb/MK cells but not in NIH/3T3 fibroblasts. Together these results indicate that Balb/MK keratinocytes possess high affinity KGF receptors to which the FGFs may also bind. However, these receptors are distinct from the receptor(s) for aFGF and bFGF on NIH/3T3 fibroblasts, which fail to interact with KGF.     

Elastic network models for RNA: a comparative assessment with molecular dynamics and SHAPE experiments

Elastic network models (ENMs) are valuable and efficient tools for characterizing the collective internal dynamics of proteins based on the knowledge of their native structures. The increasing evidence that the biological functionality of RNAs is often linked to their innate internal motions poses the question of whether ENM approaches can be successfully extended to this class of biomolecules. This issue is tackled here by considering various families of elastic networks of increasing complexity applied to a representative set of RNAs. The fluctuations predicted by the alternative ENMs are stringently validated by comparison against extensive molecular dynamics simulations and SHAPE experiments. We find that simulations and experimental data are systematically best reproduced by either an all-atom or a three-beads-per-nucleotide representation (sugar-base-phosphate), with the latter arguably providing the best balance of accuracy and computational complexity.     

Spatial distribution of extractable organohalogens in northern pink shrimp in the North Atlantic

Extractable organohalogens (EOX) are organic compounds that contain chlorine, bromine and/or iodine, which can be separated from the matrix by liquid/liquid or liquid/solid extraction. A combination of instrumental neutron activation analysis (INAA) and solvent extraction methods has been developed for the determination of EOX from the shrimpPandalus borealis. Levels of EOX were evaluated for spatial trends for shrimp caught in several areas off the Labrador coast, off the coast of Nova Scotia, and off the coast of Maine. Muscle contained 1.09–6.05 Μg EOCl/g tissue and 105–498 Μg extractable organochlorine (EOCl)/g lipid; 0.0607–0.288 Μg extractable organobromine (EOB)r/g tissue and 4.74-10.5 Μg EOBr/g lipid; and 0.014–0.048 Μg extractable organoiodine (EOI)/g tissue and 1.03–1.76 Μg EOI/g lipid, respectively. The levels of EOC1 in roe were 1.60–12.34 Μg/g tissue and 39.0-146 Μg/g lipid. In roe, the EOBr levels were 0.707–1.03 Μg/g tissue and 6.96–13.5 Μg/g lipid; and EOI levels were 0.123–0.349 Μg/g tissue and 1.42–4.11 Μg/g lipid. The EOCl, EOBr, and EOI levels in roe increased noticeably from north to south along the coast of Labrador. Samples taken from the coast of Maine and from Canso Hole were typically higher in EOCl levels than those taken from Labrador. The results for EOBr and EOI were in the same range as those from Labrador.     

The Acidic Domain and First Immunoglobulin-Like Loop of Fibroblast Growth Factor Receptor 2 Modulate Downstream Signaling through Glycosaminoglycan Modification

Fibroblast growth factor receptors (FGFRs) are membrane-spanning tyrosine kinases that have been implicated in a variety of biological processes including mitogenesis, cell migration, development, and differentiation. We identified a unique isoform of FGFR2 expressed as a diffuse band with an unusually large molecular mass. This receptor is modified by glycosaminoglycan at a Ser residue located immediately N terminal to the acidic box, a stretch of acidic amino acids. The acidic box and the glycosaminoglycan modification site are encoded by an alternative exon of the FGFR2 gene. The acidic box appears to play an important role in glycosaminoglycan modification, and the presence of this domain is required for modification by heparan sulfate glycosaminoglycan. Moreover, the presence of the first immunoglobulin-like domain encoded by another alternative exon abrogated the modification. The high-affinity receptor with heparan sulfate modification enhanced receptor autophosphorylation, substrate phosphorylation, and ternary complex factor-independent gene expression. It also sustained mitogen-activated protein kinase activity and increased eventual DNA synthesis, a long-term response to fibroblast growth factor stimulation, at physiological ligand concentrations. We propose a novel regulation mechanism of FGFR2 signal transduction through glycosaminoglycan modification.     

Stimulation of growth hormone release by thyrotropin-releasing hormone in elderly subjects

The effect of thyrotropin-releasing hormone (TRH) on the release of growth hormone (GH) was investigated in 16 elderly male subjects aged 74-88 years. Intravenous injection of 200 micrograms TRH induced a clear-cut GH rise (greater than or equal to 10 ng/ml) in 7 of 16 subjects. TRH administration did not raise plasma GH in 10 adult subjects aged 36-58 years. The results suggest disorders in neurobiochemical mechanisms regulating hypothalamopituitary function in elderly men.     

Pulsed-field electrophoresis screening for immunoglobulin heavy-chain constant-region (IGHC) multigene deletions and duplications.

Genome regions containing multiple copies of homologous genes, such as the immunoglobulin (Ig) heavy-chain constant-region (IGHC) locus, are often unstable and give rise to duplicated and deleted haplotypes. Analysis of such processes is fundamental to understanding the mechanisms of evolution of multigene families. In the IGHC region, a number of single and multiple gene deletions, derived from either unequal crossing-over or looping-out excision, have been described. To study these haplotypes at the population level, a simple and efficient method for preparing large numbers of DNA samples suitable for pulsed-field gel electrophoresis (PFGE) analysis was set up, and a sample of 110 blood donors was screened. Deletions were found to be frequent, as expected on the basis of previous serological surveys for homozygotes. Furthermore, a number of multigene duplications, never identified before, were detected. The total frequency of individuals bearing rearranged IGHC haplotypes was 10%. The genes involved in these deletions and duplications were assessed by densitometric analysis of standard Southern blots hybridized with several IGHC probes; two types of deletion and two types of duplication could thus be characterized. These data provide further evidence of the instability of the IGHC locus and demonstrate that unequal crossing-over is the most likely origin of rearranged IGHC haplotypes; they also suggest that such recombination events may be relatively frequent. Moreover, the simplicity and effectiveness of the large-scale PFGE screening approach will be of great help in the study of multigene families and of other loci involved in aberrant recombinations.     

CD23<Superscript>+</Superscript>/CD21<Superscript>hi</Superscript> B-cell translocation and ipsilateral lymph node collapse is associated with asymmetric arthritic flare in TNF-Tg mice

Introduction  

Rheumatoid arthritis (RA) is a chronic autoimmune disease with episodic flares in affected joints. However, how arthritic flare occurs only in select joints during a systemic autoimmune disease remains an enigma. To better understand these observations, we developed longitudinal imaging outcomes of synovitis and lymphatic flow in mouse models of RA, and identified that asymmetric knee flare is associated with ipsilateral popliteal lymph node (PLN) collapse and the translocation of CD23⁺/CD21^hi B-cells (B-in) into the paracortical sinus space of the node. In order to understand the relationship between this B-in translocation and lymph drainage from flaring joints, we tested the hypothesis that asymmetric tumor necrosis factor (TNF)-induced knee arthritis is associated with ipsilateral PLN and iliac lymph node (ILN) collapse, B-in translocation, and decreased afferent lymphatic flow.