bluuebunny/crossref_metadata_2025 huggingface dataset:
Amyloid plaque core protein in Alzheimer disease and Down syndrome.
Authors: Masters C L , Simms G , Weinman N A , Multhaup G , McDonald B L , Beyreuther K
Published: May 2006

We have purified and characterized the cerebral amyloid protein that forms the plaque core in Alzheimer disease and in aged individuals with Down syndrome. The protein consists of multimeric aggregates of a polypeptide of about 40 residues (4 kDa). The amino acid composition, molecular mass, and NH2-terminal sequence of this amyloid protein are almost identical to those described for the amyloid deposited in the congophilic angiopathy of Alzheimer disease and Down syndrome, but the plaque core proteins have ragged NH2 termini. The shared 4-kDa subunit indicates a common origin for the amyloids of the plaque core and of the congophilic angiopathy. There are superficial resemblances between the solubility characteristics of the plaque core and some of the properties of scrapie infectivity, but there are no similarities in amino acid sequences between the plaque core and scrapie polypeptides.
Aberrant neurofilament phosphorylation in Alzheimer disease.
Authors: Sternberger N H , Sternberger L A , Ulrich J
Published: May 2006

Alzheimer tangles, despite their location in neuronal perikarya, react immunocytochemically with monoclonal antibodies to phosphorylated epitopes of neurofilaments. Normal perikarya do not contain phosphorylated neurofilaments. The aberrant phosphorylation in both plaques and tangles seems to be largely restricted to individual phosphorylation sites among the many sites available in neurofilaments. It is suggested that the Alzheimer lesion involves an imbalance within specific kinases responsible for phosphorylation of different sites in neurofilaments.
Peptides from regenerating central nervous system promote specific populations of macroglia.
Authors: Giulian D , Tomozawa Y , Hindman H , Allen R L
Published: May 2006

The regenerating central nervous system of goldfish contains peptides referred to as glia-promoting factors (GPFs) that stimulate the proliferation of mammalian macroglia. We find that, in vitro, GPF1 and GPF3 promote the appearance of oligodendroglia and GPF2 and GPF4 stimulate proliferation of astroglia. The activities of GPF1, GPF3, and GPF4 increase during regeneration of the goldfish visual system. These results suggest that brain peptides may play a role in the recovery of the injured central nervous system by regulating the growth and development of specific macroglial populations.
Cloning, characterization, and sequence of the yeast DNA topoisomerase I gene.
Authors: Thrash C , Bankier A T , Barrell B G , Sternglanz R
Published: May 2006

The structural gene for yeast DNA topoisomerase I (TOP1) has been cloned from two yeast genomic plasmid banks. Integration of a plasmid carrying the gene into the chromosome and subsequent genetic mapping shows that TOP1 is identical to the gene previously called MAK1. Seven top1 (mak1) mutants including gene disruptions are viable, demonstrating that DNA topoisomerase I is not essential for viability in yeast. A 3787-base-pair DNA fragment including the gene has been sequenced. The protein predicted from the DNA sequence has 769 amino acids and a molecular weight of 90,020.
Faithful cell-cycle regulation of a recombinant mouse histone H4 gene is controlled by sequences in the 3'-terminal part of the gene.
Authors: Lüscher B , Stauber C , Schindler R , Schümperli D
Published: May 2006

We have analyzed the expression of endogenous histone H4 genes and of a newly introduced H4 gene in 21-Tb cells, a mouse mastocytoma cell-cycle mutant. Endogenous H4 mRNAs were less abundant by a factor of 120-180 in G1-arrested than in exponentially multiplying cells. However, H4 transcription rates were only decreased by a factor of 3 under these conditions, as determined by in vitro elongation of nascent transcripts. This indicates that post-transcriptional control of histone mRNA levels is important, in accord with published data. We introduced a mouse H4 gene, modified by a 12-base-pair (bp) insertion in its coding sequence, into 21-Tb cells by DNA-mediated gene transfer. The levels of transcripts from this gene were regulated in parallel with those of the endogenous genes. Moreover, fusion of the simian virus 40 (SV40) early promoter to a 463-bp fragment containing the 3'-terminal half of the mouse H4 gene, including 230 bp of spacer sequences, led to the regulated expression of SV40/H4 fusion RNA. However, a small proportion of SV40-initiated transcripts were not processed to histone-specific 3' ends, but extended farther through the downstream Escherichia coli galactokinase gene to a SV40 polyadenylylation site. In contrast to the short SV40/H4 RNA, the levels of these longer transcripts were not reduced in G1-arrested cells. These results show that sequences in the 3'-terminal part of the H4 gene can regulate gene expression in the cell cycle, presumably at the post-transcriptional level, as long as they are not positioned much more distant from the terminus than normal.
Lysosomal enzyme binding to mouse P388D1 macrophage membranes lacking the 215-kDa mannose 6-phosphate receptor: evidence for the existence of a second mannose 6-phosphate receptor.
Authors: Hoflack B , Kornfeld S
Published: May 2006

Mouse P388D1 macrophages target newly synthesized acid hydrolases to lysosomes in spite of their lack of the 215-kDa mannose 6-phosphate (Man-6-P) receptor. We now report that these cells contain a membrane-associated Man-6-P receptor that is distinct from the previously described receptor. The new receptor binds lysosomal enzymes containing phosphomannosyl residues. This binding is inhibited by Man-6-P or by pretreatment of the lysosomal enzymes with alkaline phosphatase. Lysosomal enzyme binding occurs at neutral pH and dissociation of the bound ligand occurs at low pH values comparable to those found within endosomes or lysosomes. The new receptor differs from the 215-kDa Man-6-P receptor in two ways. It has an absolute requirement for divalent cations and is unable to bind Dictyostelium discoideum lysosomal enzymes, which contain methylphosphomannosyl residues rather than the usual phosphomannosyl monoesters. Based on the difference in cation requirement, we suggest that the 215-kDa receptor be referred to as Man-6-P receptor CI (cation independent) and the new receptor as Man-6-P receptor CD (cation dependent). We conclude that the Man-6-P receptor CD functions in the targeting of newly synthesized acid hydrolases to lysosomes in P388D1 macrophages.
Hepatitis B virus integration in hepatocellular carcinoma DNA: duplication of cellular flanking sequences at the integration site.
Authors: Yaginuma K , Kobayashi M , Yoshida E , Koike K
Published: May 2006

The integrated form of hepatitis B virus (HBV) in the human hepatoma cell line huH2-2 and its cellular counterpart sequence have been cloned and analyzed. Blot hybridization analysis and nucleotide sequencing indicated that a single copy of the 1895-base-pair (bp) subgenomic region of HBV DNA, spanning from the middle of pre-S to the end of gene X, was integrated and flanked by the 12-bp directly repeating cellular sequences. A comparison of the sequencing data with that of the cellular counterpart DNA indicated the absence of deletion and rearrangement in the cellular flanking DNA following integration of the 1895-bp HBV DNA, except for generation of the 12-bp direct repeat at the virus-cell junction. A possible model for the mechanism of HBV integration is proposed.
Influence of dendritic cells on tumor growth.
Authors: Knight S C , Hunt R , Dore C , Medawar P B
Published: May 2006

Dendritic cells (DC) exposed to antigen are potent initiators of immune responses, and the numbers of DC and the dose of antigen control the level of response. The influence of these variables was tested on the growth of mouse sarcoma cells in vivo. When normal syngeneic DC (100,000) were given to mice with palpable tumors, tumor regression or delay in tumor growth was obtained. DC exposed to increasing doses of tumor extract in vitro before administration had progressively less effect. DC exposed to antigen delayed tumor growth significantly only when given on the same day as 500 tumor cells. The studies suggested that low doses of antigen on DC elicit immune responses and that high doses block them. The numbers of antigen-presenting cells and the dose of antigen modulate the degree of immunity to mouse sarcoma in vivo.
The nucleotide sequence of the gene for human protein C.
Authors: Foster D C , Yoshitake S , Davie E W
Published: May 2006

A human genomic DNA library was screened for the gene for protein C by using a cDNA probe coding for the human protein. Three different overlapping lambda Charon 4A phage were isolated that contain inserts for the gene for protein C. The complete sequence of the gene was determined by the dideoxy method and shown to span about 11 kilobases of DNA. The coding and 3' noncoding portion of the gene consists of eight exons and seven introns. The eight exons code for a preproleader sequence of 42 amino acids, a light chain of 155 amino acids, a connecting dipeptide of Lys-Arg, and a heavy chain of 262 amino acids. The preproleader sequence and the connecting dipeptide are removed during processing, resulting in the mature protein composed of a heavy and a light chain held together by a disulfide bond. The heavy chain also contains the catalytic region for the serine protease. Two Alu sequences and two homologous repeats of about 160 nucleotides were found in intron E. The seven introns in the gene for protein C are located in essentially the same positions in the amino acid sequence as the seven introns in the gene for human factor IX, while the first three introns in protein C are located in the same positions as the first three in the gene for human prothrombin.
Group translocation as a mechanism for sucrose transfer into vacuoles from sugarcane cells
Authors: Thom Margaret , Maretzki Andrew
Published: May 2006

Isolated vacuoles from sugarcane cells took up uridine diphosphate glucose (UDP-Glc) from the surrounding medium at a rapid rate. After a 7-min incubation of vacuoles with UDP-[14C]Glc, sucrose and sucrose phosphate were identified in the vacuole extract. UDP-Glc in the incubation medium was converted to hexose phosphates, sucrose, and glucose, with very little UDP-Glc remaining. Fructose 6-phosphate was not required for UDP-Glc uptake nor was [
14
C]fructose 6-phosphate taken up even in the presence of UDP-Glc. Glucose 6-phosphate and glucose 1-phosphate also were not taken up into vacuoles. UDP-Glc uptake showed saturation kinetics with a
K
m
of 0.7 mM and a
V
max
of 11.1 nmol/min per 10
6
vacuoles. The optimum pH for UDP-Glc uptake was between 6.5 and 7.0. Uptake of UDP-Glc could be inhibited by
p
-chloromercuriphenylsulfonic acid, UDP, and GDP, and to a lesser extent by carbonyl cyanide
m
-chlorophenylhydrazone. The UDP-Glc binding site was specific for UDP-Glc; adenosine diphosphate glucose was not taken up, and guanosine diphosphate glucose did not compete with UDP-Glc for the binding site. The results suggest that sucrose transfer into vacuoles from sugarcane is via a group translocation mechanism, probably involving five tonoplast-bound enzymes.
Immunochemical characterization of carboxypeptidase B-like peptide-hormone-processing enzyme.
Authors: Hook V Y , Mezey E , Fricker L D , Pruss R M , Siegel R E , Brownstein M J
Published: May 2006

Specific rabbit antisera against purified bovine pituitary carboxypeptidase processing enzyme (which has also been referred to as "enkephalin convertase") have been prepared and characterized. The antisera recognized both the purified soluble and the membrane-bound forms of the enzyme with equal affinity, suggesting that these two forms of the enzyme may possess many regions of structural homology. Since the antisera did not crossreact with carboxypeptidases B, N, A, Y, and P, the carboxypeptidase processing enzyme may be a structurally distinct form of carboxypeptidase. Carboxypeptidase immunostaining, as seen by light microscopy, was found throughout the rat brain and in bovine adrenal medulla, reflecting the widespread distribution of neuropeptides. Electron microscopic immunocytochemistry of rat paraventricular nucleus and other brain areas showed that the enzyme was present in some dendrites and nerve terminals, which contain storage vesicles. These findings support the hypothesis that this carboxypeptidase is involved in the processing of many peptide hormone precursors.
Gene repertoire of the anti-poly(Glu60Ala30Tyr10) (GAT) immune response: comparison of VH, V kappa, and D regions used by anti-GAT antibodies and monoclonal antibodies produced after anti-idiotypic immunization.
Authors: Roth C , Rocca-Serra J , Sommé G , Fougereau M , Théze J
Published: May 2006

Eight monoclonal antibodies were selected from BALB/c mice immunized with two different monoclonal anti-idiotypic antibodies recognizing two discrete idiotopes characteristic of the anti-poly(Glu60Ala30Tyr10) (GAT) antibody response. These monoclonal antibodies were previously classified as Ab1 (anti-GAT-like) and Ab3 (anti-anti-idiotype) on the basis of expression of the public idiotypic specificity (p.GAT) studied with a xenogeneic serum, anti-GAT activity, and expression of various public idiotopes. All the heavy chain variable region (VH) sequences from Ab1 are nearly identical to the VH sequences of Ab1 anti-GAT monoclonal antibodies. The same type of results has been found with the Ab1 kappa light chain variable region (V kappa) sequences. Confirming our classification, Ab3 VH and V kappa sequences were found to be completely different from Ab1 VH and V kappa sequences. The Ab1 diversity (D) regions are different from one another and different from the D regions found on monoclonal anti-GAT antibodies but function similarly. These D regions are not simply derived from already described D genes. Finally, our results suggest that in the anti-GAT response VH and V kappa sequence are mainly responsible for idiotype expression.
A glycoprotein associated with the non-A, non-B hepatitis agent(s): isolation and immunoreactivity.
Authors: Seto B , Gerety R J
Published: May 2006

A glycoprotein was isolated and purified to homogeneity from the serum of a patient with chronic non-A, non-B hepatitis. NaDodSO4/PAGE of the glycoprotein revealed a single major band at Mr approximately 77,000. Antibodies to this glycoprotein were shown to possess the following immunoreactivity: (i) they reacted by radioimmunoassay with sera obtained at the time of diagnosis from 17 of 42 patients with non-A, non-B hepatitis and with only 2 of 58 sera from either matched controls or patients with hepatitis A or hepatitis B, (ii) they reacted with sucrose gradient fractions from a proven infectious non-A, non-B hepatitis serum at a peak density of 1.14 g/ml and in the soluble protein fractions on top of the gradient, and (iii) they reacted in ELISA with disrupted human T-cell lymphocytotropic virus type III (HTLV-III), and (iv) they reacted in immunoblots with a protein of Mr 74,000 derived from HTLV-III.
Gametophytic self-incompatibility is controlled by a single major locus on chromosome 1 in
Lycopersicon peruvianum
Authors: Tanksley Steven D. , Loaiza-Figueroa Fernando
Published: May 2006

By using a number of previously mapped enzyme-coding genes as genetic markers, it has been possible to scan the genome of
Lycopersicon peruvianum
for gene(s) controlling the gametophytic self-incompatibility reaction. Regardless of genetic background or level of inbreeding, only a single locus (
S
), mapping to chromosome 1, was found to control the self-incompatibility reaction. Despite the widespread occurrence of this form of self-incompatibility in higher plants, to the best of our knowledge, the locus underlying the response has not been confirmed previously through genetic mapping, and the results cast doubts on hypotheses requiring multifactoral or dynamic control of gametophytic self-incompatibility.
Pulse-wave propagation in an artery with leakage into small side branches.
Authors: Chadwick R S
Published: May 2006

Linear pulse propagation theory is applied to a network of elastic tubes. The network is specialized to one having a main branch with many low-admittance side branches. Continuum approximations and an asymptotic analysis of side-branch network input admittance reduce the original network to a tapered leaky tube. The outflow law is equivalent to distributed Windkessels. The conductance and time constant of the Windkessels are related to the geometry of the side-branch networks. Viscous effects are included in the theory. Asymptotic methods are used to determine the dispersion relation and waveform amplitude in the frequency domain. Periodic pressure and flow waveforms are computed using Fourier synthesis.
Identical short peptide sequences in unrelated proteins can have different conformations: a testing ground for theories of immune recognition.
Authors: Wilson I A , Haft D H , Getzoff E D , Tainer J A , Lerner R A , Brenner S
Published: May 2006

The ability of antibodies raised against disordered short peptides to interact frequently with their cognate sequences in intact folded proteins has raised a major theoretical issue in protein chemistry. We propose to address this issue by using antibodies raised against peptides with identical sequences, but different conformations, in pairs of unrelated proteins of known three-dimensional structure. The general search method presented here enabled us to detect candidate sequences for such immunological studies.
Expression of cell-cycle-dependent genes in phytohemagglutinin-stimulated human lymphocytes.
Authors: Kaczmarek L , Calabretta B , Baserga R
Published: May 2006

We have investigated the expression of certain cell-cycle-dependent genes in human peripheral blood mononuclear cells (PBMC) stimulated by phytohemagglutinin (PHA). The genes studied had been previously identified as cell-cycle dependent in other cell types from different species and were induced by different mitogens. One of these genes (2F1) and the gene for the interleukin 2 receptor were induced by PHA even in cultures partially depleted of accessory cells where the lymphocytes grew in size but failed to enter S phase. The other genes (c-myc, 4F1, JE-3, and KC-1) were induced only in complete cultures of PBMC stimulated by PHA. These results confirm the dissociation between growth in size and cell DNA replication that can occur during cell-cycle progression. Moreover, the time course of appearance of detectable levels of RNA for these genes suggests that they may be used as markers of cell-cycle progression in the transition of lymphocytes from G0 to S phase.
Chromosomal radiosensitivity during the G2 cell-cycle period of skin fibroblasts from individuals with familial cancer.
Authors: Parshad R , Sanford K K , Jones G M
Published: May 2006

We reported previously that human cells after neoplastic transformation in culture had acquired an increased susceptibility to chromatid damage induced by x-irradiation during the G2 phase of the cell cycle. Evidence suggested that this results from deficient DNA repair during G2 phase. Cells derived from human tumors also showed enhanced G2-phase chromosomal radiosensitivity. Furthermore, skin fibroblasts from individuals with genetic diseases predisposing to a high risk of cancer, including ataxia-telangiectasia, Bloom syndrome, Fanconi anemia, and xeroderma pigmentosum exhibited enhanced G2-phase chromosomal radiosensitivity. The present study shows that apparently normal skin fibroblasts from individuals with familial cancer--i.e., from families with a history of neoplastic disease--also exhibit enhanced G2-phase chromosomal radiosensitivity. This radiosensitivity appears, therefore, to be associated with both a genetic predisposition to cancer and a malignant neoplastic state. Furthermore, enhanced G2-phase chromosomal radiosensitivity may provide the basis for an assay to detect genetic susceptibility to cancer.
Induction of experimental allergic encephalomyelitis in Lewis rats with purified synthetic peptides: delineation of antigenic determinants for encephalitogenicity, in vitro activation of cellular transfer, and proliferation of lymphocytes.
Authors: Mannie M D , Paterson P Y , U'Prichard D C , Flouret G
Published: May 2006

Four highly purified synthetic peptides encompassing segments of the 68-86 region [for the numbering system used, see Eylar, E.H., Brostoff, S., Hashim, G., Caccam, J. & Burnett, P. (1971) J. Biol. Chem. 246, 5770-5784] of myelin basic protein (MBP), a region known to induce experimental allergic encephalomyelitis (EAE) in Lewis rats, were used to define and compare structure-function relationships between the primary structure of the 68-86 sequence and the three following biological activities: induction of EAE in Lewis rats, stimulation of T lymphocytes in vitro as measured by augmented cellular transfer of EAE to syngeneic recipients, and lymphocyte proliferation, as measured by [3]thymidine incorporation. Guinea pig (GP) MBP was approximately 60 or 1500 times more active than the GP68-84 (Y G S L P Q K S Q R S Q D E N; single-letter amino acid abbreviations) or the modified bovine (MB) 68-84 (Y G S L P Q K A Q R P Q D E N) peptides for induction of EAE, respectively. Furthermore, lymphocytes primed with either GPMBP, GP68-84, or MB68-84 crossreacted in vitro with either GPMBP, GP68-84, or MB68-84 for activation of lymphocyte transfer activity. In contrast, lymphocytes primed with either GP68-84 or MB68-84 exhibited antigen-specific proliferation in vitro exclusively in response to either GP or MB sequences, respectively. Neither GP75-84 (S Q R S Q D E N) nor GP75-86 (S Q R S Q D E N P V) induced EAE, activated lymphocytes for EAE transfer, or stimulated lymphocyte proliferation under conditions and doses tested. We conclude that (i) structurally distinct determinants, reflecting existence of functionally independent classes of antigen receptors, specify encephalitogenic and proliferative responses of primed lymphocytes and (ii) determinants for EAE induction, cellular transfer of EAE, and lymphocyte proliferation include amino acid residues in the 68-74 (Y G S L P Q K) sequence of GPMBP.
Vasopressin mRNA in situ hybridization: localization and regulation studied with oligonucleotide cDNA probes in normal and Brattleboro rat hypothalamus.
Authors: Uhl G R , Zingg H H , Habener J F
Published: May 2006

Hybridizable vasopressin mRNA may be quantitatively localized in situ in sections from rat hypothalamus. Radiolabeled oligonucleotide cDNA probes, synthesized by chemical and enzymatic means, provide strong hybridization in zones known to contain vasopressin cell bodies. Multiple single-stranded 32P-, 35S-, or 3H-labeled oligonucleotides demonstrate localized hybridization that increases as probes are lengthened from 8 to 75 bases. Competition studies, RNase experiments, anatomic localization, and use of multiple probes all support hybridization specificity. An approximate doubling of hybridizable mRNA in both supraoptic and paraventricular nuclei can be detected with dehydration of the animals. Hybridizable mRNA densities are virtually normal in hypothalamic nuclei of Brattleboro rats given free access to water. These methods can provide insight into regional mRNA dynamics and may reflect functional activity of peptidergic neurons.
Experimental transmission of a microsporidian pathogen from mosquitoes to an alternate copepod host.
Authors: Andreadis T G
Published: May 2006

Meiospores of a microsporidian parasite Amblyospora sp. (Protozoa: Microspora) from larval Aedes cantator mosquitoes were directly infectious to an alternate copepod host, Acanthocyclops vernalis (Arthropoda: Crustacea). Infections ranged from 6.7% to 60.0% in laboratory tests when meiospores and copepods were maintained together for 10-30 days in filtered water from the breeding site or in a balanced salt solution. Pathogen development takes place within host adipose tissue and is fatal to the copepod. The entire developmental sequence of this microsporidian in the copepod is unikaryotic and there is no ultrastructural evidence of a sexual cycle or a restoration of the diploid condition in the alternate host. Single uninucleated spores similar to those previously described for the genus Pyrotheca are formed. Results demonstrate that haploid meiospores of Amblyospora from mosquitoes have the function of transmitting the pathogen to another host and that members of this genus are polymorphic and have at least three distinct developmental cycles, each producing a different spore.
An invertible element of DNA controls phase variation of type 1 fimbriae of Escherichia coli.
Authors: Abraham J M , Freitag C S , Clements J R , Eisenstein B I
Published: May 2006

The expression of type 1 fimbriae (pili) of Escherichia coli is turned on and off at the transcriptional level at a high frequency (10(-3) per cell per generation) in a process termed phase variation. Using Southern blot and DNA sequence analysis, we have detected a genomic rearrangement in the switch region immediately upstream of the fimbrial structural gene. This rearrangement involves an invertible 314-base-pair segment of DNA whose alternating orientation apparently results in the on-and-off activation of a promoter that determines the state of fimbrial expression.
beta-Endorphin: surface binding and internalization in thymoma cells.
Authors: Schweigerer L , Schmidt W , Teschemacher H , Gramsch C
Published: May 2006

The opioid peptide beta-endorphin binds to specific nonopioid binding sites (Mr 72,000) that are present on the surface of thymoma cells. beta-Endorphin is then internalized, apparently via the Mr 72,000 species, and is subsequently found within intracellular, vesicular structures. This process is accompanied by the down-regulation of the Mr 72,000 binding sites. Our findings suggest that beta-endorphin may modulate cellular functions, such as T-lymphocyte proliferation, at intracellular rather than cell surface sites.
Cellular receptor for 125I-labeled tumor necrosis factor: specific binding, affinity labeling, and relationship to sensitivity.
Authors: Kull F C , Jacobs S , Cuatrecasas P
Published: May 2006

Tumor necrosis factor (TNF) is a proteinaceous toxin shed by stimulated myeloid cells. Murine TNF was radioiodinated to a specific activity of 1 mCi/nmol (1 Ci = 37 GBq) of monomer. 125I-labeled TNF (125I-TNF) retained complete cytotoxic activity and it was immunochemically identical to the native toxin in a quantitative immunoprecipitation assay. It could be shown by competition binding that 125I-TNF bound to intact L929 cells with a specificity equal to that of native toxin. The conditions of time, temperature, and concentration involved in equilibrium specific binding to intact cells were studied in detail. When binding was carried out at 4 degrees C for 18 hr, four cell lines sensitive to 125I-TNF cytotoxicity demonstrated high-affinity binding. The binding reached half-maximal level at 3 pM and saturated at 30 pM. These concentrations approximated those required for cell death. Scatchard analysis gave approximately 1000 sites per cell. J774.1 cells, the source of the toxin, demonstrated similar binding but were not sensitive to 125I-TNF cytotoxicity. Other sensitive cell lines and freshly extracted tumor cells showed specific binding at 3 pM. Normal lymphoid organ cell suspensions and two human tumorigenic cell lines were not sensitive and failed to demonstrate specific binding. 125I-TNF, covalently cross-linked to its receptor on sensitive L-M cells with disuccinimidyl suberate, was isolated and analyzed by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and autoradiography. Two specific bands were identified. The most prominent band had a mobility corresponding to a molecular mass of 95 kDa and the second band had a molecular mass of 75 kDa. The presence of the binding site appears to be necessary but not sufficient to explain the sensitivity of cells to the cytotoxic action of TNF.
Comparative statistics for DNA and protein sequences: single sequence analysis.
Authors: Karlin S , Ghandour G
Published: May 2006

Four categories of data representations are used to help interpret structures and similarities of nucleic acid and protein sequences. Statistical significance of the observed relationships revealed by these representations are assessed by a hierarchy of permutation procedures and by comparisons with theoretical random models. Applications are presented for various DNA sequences including papovaviruses, Epstein-Barr virus, mitochondrial genomes, and several globin and immunoglobulin genes.
Evidence for the involvement of the submandibular gland epidermal growth factor in mouse mammary tumorigenesis.
Authors: Kurachi H , Okamoto S , Oka T
Published: May 2006

The submandibular gland is a rich source of epidermal growth factor (EGF) in mice. The concentration of EGF in the gland of virgin female mice of C3H/HeN strain increased as much as 9-fold from the age of 30 to 52 weeks. During this period, the incidence of mammary tumor in virgin females increased markedly to a maximal level of 62.5% (n = 48) at 52 weeks of age. Removal of the submandibular gland (sialoadenectomy) of virgin mice 14-22 weeks old reduced the tumor incidence to 12.8% (n = 39) at the age of 52 weeks and also increased the latency period of mammary tumor development as much as 14 weeks when compared to that of normal mice. Long-term treatment of sialoadenectomized virgin mice with EGF (5 micrograms per mouse every other day) increased the tumor incidence to 33.3%. Moreover, sialoadenectomy of mammary tumor-bearing animals caused a rapid and sustained cessation of tumor growth, but EGF administration (5 micrograms per mouse per day) quickly restored the rate of tumor growth to a normal level. These results indicate that submandibular gland EGF plays a crucial role in mouse mammary tumorigenesis.
Endothelial cell-derived chemotactic activity for mouse peritoneal macrophages and the effects of modified forms of low density lipoprotein.
Authors: Quinn M T , Parthasarathy S , Steinberg D
Published: May 2006

Cultured rabbit and bovine aortic endothelial cells generated chemotactic activity for mouse resident peritoneal macrophages, demonstrable in the conditioned medium. This chemotactic activity was heat stable and was not extracted into chloroform/methanol. It was inhibited by addition of endothelial cell-modified low density lipoprotein (EC-modified LDL), a form of LDL shown previously to contain peroxidized lipids, increased lysophosphatidylcholine, and partially degraded apoprotein B. The chemotactic activity was also inhibited by LDL previously oxidized in the absence of cells with 5 microM Cu2+. Inhibitory activity was present in the lipid extract of EC-modified LDL but not in that of native LDL, presumably representing peroxidized lipid components. EC-modified LDL also inhibited the chemotactic activity of zymosan-activated serum. Because EC-modified LDL is taken up in part by way of the acetyl-LDL receptor, the effects of acetyl-LDL were tested. Rather than inhibiting chemotaxis, acetyl LDL showed intrinsic positive chemotactic activity as did also fucoidin and polyinosinic acid, both of which also interact with the acetyl-LDL receptor. These studies suggest mechanisms by which macrophages may be recruited into the subendothelial space by endothelial cell-derived chemotactic factors or by natural polyanions structurally related to fucoidin or polyinosinic acid and then become "trapped" there because of the inhibitory effects of peroxidized lipid components in modified forms of LDL.
Predominance of hemodynamic rather than metabolic factors in the pathogenesis of diabetic glomerulopathy.
Authors: Zatz R , Meyer T W , Rennke H G , Brenner B M
Published: May 2006

Six groups of Munich-Wistar rats underwent micropuncture study 2-10 weeks and morphologic studies 11-13 months after induction of streptozotocin diabetes or after sham treatment. Diabetic rats received diets containing 6% (group D6), 12% (D12), or 50% protein (D50) and were maintained under similar conditions of moderate hyperglycemia by daily injections of ultralente insulin. Age- and weight-matched normal control rats were also given 6% (Group N6), 12% (N12), or 50% protein (N50). Kidney weight, whole-kidney and single-nephron glomerular filtration rate, glomerular plasma flow, and mean glomerular transcapillary hydraulic pressure difference were higher in D50 rats than in all other groups and predisposed this group to marked and progressive albuminuria. Likewise, histological examination of the kidneys disclosed areas of sclerosis in 19.6% of glomeruli in D50 rats; the frequency of such lesions was less than 2.5% in all other groups. These findings indicate that the metabolic disorder seen in stable, moderately hyperglycemic diabetic rats does not lead to glomerulopathy as long as elevations in glomerular pressures and flows are prevented.
Differential expression of distinct microtubule-associated proteins during brain development.
Authors: Riederer B , Matus A
Published: May 2006

The levels of three different microtubule-associated proteins (MAP1, -2, and -3) in brain were found to undergo large changes during postnatal development. MAP1 was barely detectable at birth but thereafter steadily increased, reaching adult levels by postnatal day 20 (P20). Both MAP2 and MAP3 showed differential expression patterns of their component peptides. At birth, MAP2 was represented by the smaller of two Mr 280,000 peptides (MAP2b) and three antigenically related Mr 70,000 peptides. The larger of the Mr 280,000 peptides (MAP2a) first appeared between P10 and P20, and the Mr 70,000 components disappeared at the same time. Of the two MAP3 peptides, the larger (MAP3a) was present in the late embryo, several days before MAP3b appeared. Between P10 and P20, both MAP3 components underwent a striking decrease in abundance (a factor of 10), which correlated with their disappearance from all neuronal compartments except neurofilament-containing axons. These developmental changes in expression are different and characteristic for each of the three MAPs, yet in each case they are detectable in brain homogenates, indicating that they occur concurrently throughout the brain.
Cloning and expression in Escherichia coli of the cDNA for murine tumor necrosis factor.
Authors: Pennica D , Hayflick J S , Bringman T S , Palladino M A , Goeddel D V
Published: May 2006

A murine tumor necrosis factor (MuTNF) cDNA was isolated from a cDNA library prepared by using mRNA from the murine macrophage-like cell line PU5-1.8 induced with 4 beta-phorbol 12 beta-myristate 13 alpha-acetate. The cDNA encodes a polypeptide consisting of a 79 amino acid pre sequence followed by a mature MuTNF sequence of 156 amino acids. The 235 amino acid murine pre-TNF polypeptide is 79% homologous to the human pre-TNF protein. There is one potential N-linked glycosylation site on MuTNF, in contrast to human TNF, which lacks any such site. The MuTNF cDNA, when engineered for expression in Escherichia coli, was found to direct the synthesis of biologically active MuTNF as determined by its cytotoxicity against several transformed cell lines.
Human placental alkaline phosphatase in liver and intestine.
Authors: Garattini E , Margolis J , Heimer E , Felix A , Udenfriend S
Published: May 2006

Three distinct forms of human alkaline phosphatase, presumably isozymes, are known, each apparently associated with a specific tissue. These are placental, intestinal, and liver (kidney and bone). We have used a specific immunoassay and HPLC to show that placental alkaline phosphatase is also present in extracts of liver and intestine in appreciable amounts.
Visualization of the protein associations in the erythrocyte membrane skeleton.
Authors: Byers T J , Branton D
Published: May 2006

We have obtained clear images of the erythrocyte membrane skeleton from negatively stained preparations that originate directly from the intact cell but in which the spectrin meshwork is artificially spread to allow close inspection. Our procedure requires less than 2 min at 5 degrees C in phosphate buffers. We find 200-nm-long spectrin tetramers crosslinked by junctional complexes. Each junction contains a regular 37-nm rod, probably an actin oligomer of approximately 13 monomers. Densities appear at variable places in the meshwork but distinct globules occur with great frequency 78 nm from the spectrin tetramer's junctional insertion end, very close to the known binding site for ankyrin. Most frequently, five or six spectrin tetramers insert into each junction, producing a meshwork that displays remarkably regular long range order.
Inhibition of soluble immune response suppressor activity by growth factors.
Authors: Aune T M
Published: May 2006

Soluble immune response suppressor (SIRS), a protein of Mr 14,000, is a lymphokine produced by interferon- or concanavalin A-activated suppressor T cells and is oxidized to its activated form, SIRSox, by H2O2 produced by macrophages. SIRSox inhibits antibody secretion by B lymphocytes and cell division by normal or transformed cell lines. Effects of purified growth factors on suppression of antibody secretion were examined to determine whether any would oppose the inhibitory effects of SIRS or SIRSox. Interleukin 1 (IL-1), interleukin 2 (IL-2), and epidermal growth factor (EGF) each inhibited SIRS-mediated suppression of antibody secretion by cultured mouse spleen cells. Inhibition of SIRS activity was most effective when growth factors were added late in the culture period. IL-1, IL-2, and EGF also blocked suppression by SIRSox. However, EGF and IL-1 blocked suppression by SIRSox only when added 3-6 hr before addition of SIRSox, whereas IL-2 blocked suppression by SIRSox when added before or up to 3 hr after addition of SIRSox. Further evaluation showed that IL-2, but not EGF or IL-1, reversed inhibition of antibody secretion by SIRSox in a time- and concentration-dependent manner. With 50 units of IL-2 per 0.5-ml culture, reversal was complete within 1 hr. The ability of growth factors to interfere with inhibition of cell division by SIRSox was examined with the human B-cell leukemia RPMI-1788. This cell line binds EGF but is not known to have cell surface receptors for IL-1 or IL-2. EGF (0.3-1 ng/ml), when added to RPMI-1788 cultures 4-6 hr before SIRSox, interfered with the ability of SIRSox to inhibit cell division. Taken together, these data indicate that growth factors interfere with both the immunosuppressive and growth inhibitory properties of SIRSox in both heterogeneous and homogeneous cell populations.
The influence of mevinolin on the adrenal cortical response to corticotropin in heterozygous familial hypercholesterolemia.
Authors: Illingworth D R , Corbin D
Published: May 2006

The biosynthesis of adrenal corticosteroids in humans depends on a continuous supply of cholesterol, which can be derived from both local synthesis and receptor-mediated uptake of low density lipoproteins (LDL) from plasma. Mevinolin, an inhibitor of 3-hydroxy-3-methylglutaryl-CoA reductase [mevalonate:NAD+ oxidoreductase (CoA-acylating), EC 1.1.1.88] is an effective hypolipidemic agent in patients with heterozygous familial hypercholesterolemia. To determine whether mevinolin influences the adrenal production of corticosteroids, the adrenocortical response to a continuous 36-hr infusion of corticotropin (ACTH) was examined in eight patients with heterozygous familial hypercholesterolemia before, and again during, treatment with mevinolin (40-80 mg/day). The drug produced an average decrease of 28% and 34% in the plasma concentrations of total and LDL cholesterol. Serum cortisol levels showed similar increases in response to ACTH stimulation before and during mevinolin treatment, and the rates of excretion of urine-free cortisol were also similar. We conclude that clinically effective doses of mevinolin do not affect corticosteroid production by the adrenal cortex during prolonged ACTH stimulation in patients with heterozygous familial hypercholesterolemia.
Two types of muscarinic response to acetylcholine in mammalian cortical neurons.
Authors: McCormick D A , Prince D A
Published: May 2006

Applications of acetylcholine (AcCho) to pyramidal cells of guinea pig cingulate cortical slices maintained in vitro result in a short latency inhibition, followed by a prolonged increase in excitability. Cholinergic inhibition is mediated through the rapid excitation of interneurons that utilize the inhibitory neurotransmitter gamma-aminobutyric acid (GABA). This rapid excitation of interneurons is associated with a membrane depolarization and a decrease in neuronal input resistance. In contrast, AcCho-induced excitation of pyramidal cells is due to a direct action that produces a voltage-dependent increase in input resistance. In the experiments reported here, we investigated the possibility that these two responses are mediated by different subclasses of cholinergic receptors. The inhibitory and slow excitatory responses of pyramidal neurons were blocked by muscarinic but not by nicotinic antagonists. Pirenzepine was more effective in blocking the AcCho-induced slow depolarization than in blocking the hyperpolarization of pyramidal neurons. The two responses also varied in their sensitivity to various cholinergic agonists, making it possible to selectively activate either. These data suggest that AcCho may produce two physiologically and pharmacologically distinct muscarinic responses on neocortical neurons: slowly developing voltage-dependent depolarizations associated with an increase in input resistance in pyramidal cells and short-latency depolarizations associated with a decrease in input resistance in presumed GABAergic interneurons.
Cloning and characterization of two cDNAs coding for human von Willebrand factor.
Authors: Sadler J E , Shelton-Inloes B B , Sorace J M , Harlan J M , Titani K , Davie E W
Published: May 2006

A cDNA library was prepared in lambda gt11 bacteriophage from poly(A)+ RNA isolated from primary cultures of endothelial cells from human umbilical vein. Approximately 2.5 million independent recombinants were screened and 2 of those were found to synthesize a fusion protein with beta-galactosidase that reacted with rabbit antibody against human von Willebrand factor. Comparison of the amino acid sequence translated from the cDNA insert of the two clones with the amino acid sequence determined by Edman degradation of the protein established that both phage isolates code for von Willebrand factor. The first clone (lambda HvWF1) contained an insert of 404 nucleotides that corresponded to amino acid residues 1-110 in the mature protein circulating in blood, in addition to a portion (24 amino acids) of a prepro leader sequence. The second cDNA clone (lambda HvWF3) contained an insert of 4.9 kilobases that coded for the carboxyl-terminal 1525 amino acids of von Willebrand factor, a stop codon of TGA, 134 nucleotides of 3' noncoding sequence, and a poly(A) tail of 150 nucleotides. The two clones together code for greater than 80% of the molecule circulating in blood. The same carboxyl-terminal lysine residue was identified in the mature protein as well as in the cDNA, indicating that all of the proteolytic processing that occurs during the biosynthesis and assembly of von Willebrand factor is associated with the amino-terminal portion of the precursor protein. The amino acid sequence of von Willebrand factor indicates the presence of two different internal gene duplications and one triplication. These repetitive amino acid sequences account for about one-half of the amino acids present in the mature protein. The tetrapeptide sequence of Arg-Gly-Asp-Ser, which mediates the cell attachment and platelet binding activity of fibronectin, was also identified in the carboxyl-terminal portion of von Willebrand factor.
Identification of a nuclear localization signal of a yeast ribosomal protein.
Authors: Moreland R B , Nam H G , Hereford L M , Fried H M
Published: May 2006

To identify a signal involved in transporting a ribosomal protein to the nucleus, we constructed hybrid genes encoding amino-terminal segments of yeast ribosomal protein L3 joined to the amino-terminal end of the entire Escherichia coli beta-galactosidase molecule. The subcellular locations of the corresponding hybrid proteins in yeast were determined by in situ immunofluorescence. The first 21 amino acids of L3 were sufficient to localize beta-galactosidase to the nucleus. This region shows limited homology to portions of other nuclear proteins identified as essential for their transport. Larger fusion proteins were also localized to the nucleus. However, a hybrid protein containing all but the 14 carboxyl-terminal amino acids from L3 initially failed to localize; this defect was corrected by inserting a glycine- and proline-containing bridge between the L3 and beta-galactosidase moieties. The renovated protein was able to associate with ribosomes, suggesting that, in addition to entering the nucleus, this hybrid polypeptide was assembled into 60S ribosomal subunits that were subsequently exported to the cytoplasm.
Retroviral insertional mutagenesis of a target allele in a heterozygous murine cell line.
Authors: Frankel W , Potter T A , Rosenberg N , Lenz J , Rajan T V
Published: May 2006

A clonal murine cell line that is heterozygous at the beta 2-microglobulin locus (B2m) was obtained by Abelson murine leukemia virus (Ab-MuLV) transformation of liver cells from (C57BL/6 X BALB/c) F1 fetuses. To obtain proviral insertional mutants, we superinfected a subclone of these cells, which does not express the env surface protein of the Moloney leukemia virus (Mo-MuLV, the helper virus that was used to transmit the defective Ab-MuLV genome during transformation), with Mo-MuLV. Mutant clones that fail to express the C57BL/6 allele of B2m (B2mb) were then immunoselected by using a monoclonal antibody that specifically recognizes the B2mb gene product and not that of the B2ma allele. Of 22 independent clones obtained, one contains a proviral insertion that is near or in the first exon of the B2mb gene. Surprisingly, the insertion is of the Ab-MuLV genome and not of replication-competent Mo-MuLV. This indicates that superinfection with Mo-MuLV "rescued" the defective Ab-MuLV genome, which then inserted into the B2mb gene. We conclude that when an allele-specific selection procedure exists, proviral insertion is a potential method for obtaining mutations in heterozygous mammalian cells. This approach may thereby provide a method for molecular cloning of such selectable genes, using a retroviral hybridization probe.