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Biomedical subjects

K R Johnson

Publications and source records attributed to K R Johnson.

At least 109 records · Page 6Linked to original sources
PubMed1995

Assignment to chromosome 11 of mouse p68 RNA helicase gene (Hlr1) and pseudogene (Hlr1-ps1).

The gene encoding murine p68 RNA helicase (Hlr1) was mapped to the distal portion of mouse chromosome 11 by linkage analysis of DNA restriction length polymorphisms using an interspecific genetic backcross between (C57BL/6J x SPRET/Ei) F1 hybrids and SPRET/Ei mice. A closely related gene (Hlr1-ps1) was identified, isolated, and mapped to the proximal part of the same chromosome. Sequence analysis and PCR results suggest that Hlr1-ps1 is a pseudogene, flanked by DNA stretches similar to mouse insertion element IE118.

Animals↗
PubMed1995

Neuromuscular degeneration (nmd): a mutation on mouse chromosome 19 that causes motor neuron degeneration.

Neuromuscular degeneration, nmd, is a spontaneous autosomal recessive mutation in the mouse producing progressive hindlimb impairment caused by spinal muscular atrophy. We used an intersubspecific intercross between B6.BKs-nmd2J/+ and Mus musculus castaneus (CAST/Ei) to map the nmd mutation to mouse Chromosome (Chr) 19 with the most likely gene order: nmd-(D19Sel2, Pygm)-Cntf-Pomc2-D19Mit16-Cyp2c-Got1. nmd maps near muscle deficient, mdf, and has a very similar clinical phenotype, but allele tests and histological differences suggest that nmd is a distinct mutation at a different locus. Although closely linked, nmd recombined with the candidate genes muscle glycogen phosphorylase, Pygm, and ciliary neurotrophic factor, Cntf.

Animals↗
PubMed1995

The differential expression of N-cadherin and E-cadherin distinguishes pleural mesotheliomas from lung adenocarcinomas.

Malignant mesotheliomas are highly aggressive tumors that develop most frequently in the pleura of patients chronically exposed to asbestos. The distinction between malignant mesotheliomas and tumors of epithelial origin, particularly peripheral lung adenocarcinoma, can be difficult despite the use of immunocytochemical markers and other diagnostic tools. During embryonic development the cadherin cell-cell adhesion molecules participate in the segregation of cells into different tissues. As a result of complex mechanisms of tissue selectivity, N-cadherin is expressed by the developing pleural mesothelial cells and E-cadherin is expressed by the epithelial cells of the lung. Thus, we postulated that N-cadherin could be used as a marker of mesothelial cells and mesothelial tumors, in contrast to adenocarcinomas of the lung that are tumors of epithelial origin. We studied the expression of N-cadherin, E-cadherin and two cadherin-associated proteins, alpha-catenin and beta-catenin, in 19 pleural mesotheliomas, 16 lung adenocarcinomas and in 2 mesothelioma cell lines using specific monoclonal antibodies and immunohistochemical methods. Our results show that all mesotheliomas express high levels of N-cadherin, regardless of their histological type, in contrast to lung adenocarcinomas which expressed E-cadherin but no N-cadherin. The cadherin-associated proteins, alpha-catenin and beta-catenin, were present in both mesotheliomas and adenocarcinomas. Our results show that pleural mesotheliomas can be distinguished from lung adenocarcinomas based on the differential expression of N-cadherin and E-cadherin, using specific monoclonal antibodies and immunocytochemistry.

Adenocarcinoma↗
PubMed1995

Association of p120, a tyrosine kinase substrate, with E-cadherin/catenin complexes.

p120 was originally identified as a substrate of pp60src and several receptor tyrosine kinases, but its function is not known. Recent studies revealed that this protein shows homology to a group of proteins, beta-catenin/Armadillo and plakoglobin (gamma-catenin), which are associated with the cell adhesion molecules cadherins. In this study, we examined whether p120 is associated with E-cadherin using the human carcinoma cell line HT29, as well as other cell lines, which express both of these proteins. When proteins that copurified with E-cadherin were analyzed, not only alpha-catenin, beta-catenin, and plakoglobin but also p120 were detected. Conversely, immunoprecipitates of p120 contained E-cadherin and all the catenins, although a large subpopulation of p120 was not associated with E-cadherin. Analysis of these immunoprecipitates suggests that 20% or less of the extractable E-cadherin is associated with p120. When p120 immunoprecipitation was performed with cell lysates depleted of E-cadherin, beta-catenin was no longer coprecipitated, and the amount of plakoglobin copurified was greatly reduced. This finding suggests that there are various forms of p120 complexes, including p120/E-cadherin/beta-catenin and p120/E-cadherin/plakoglobin complexes; this association profile contrasts with the mutually exclusive association of beta-catenin and plakoglobin with cadherins. When the COOH-terminal catenin binding site was truncated from E-cadherin, not only beta-catenin but also p120 did not coprecipitate with this mutated E-cadherin. Immunocytological studies showed that p120 colocalized with E-cadherin at cell-cell contact sites, even after non-ionic detergent extraction. Treatment of cells with hepatocyte growth factor/scatter factor altered the level of tyrosine phosphorylation of p120 as well as of beta-catenin and plakoglobin. These results suggest that p120 associates with E-cadherin at its COOH-terminal region, but the mechanism for this association differs from that for the association of beta-catenin and plakoglobin with E-cadherin, and thus, that p120, whose function could be modulated by growth factors, may play a unique role in regulation of the cadherin-catenin adhesion system.

Binding Sites↗
PubMed1995

Human peroxisomal targeting signal-1 receptor restores peroxisomal protein import in cells from patients with fatal peroxisomal disorders.

Two peroxisomal targeting signals, PTS1 and PTS2, are involved in the import of proteins into the peroxisome matrix. Human patients with fatal generalized peroxisomal deficiency disorders fall into at least nine genetic complementation groups. Cells from many of these patients are deficient in the import of PTS1-containing proteins, but the causes of the protein-import defect in these patients are unknown. We have cloned and sequenced the human cDNA homologue (PTS1R) of the Pichia pastoris PAS8 gene, the PTS1 receptor (McCollum, D., E. Monosov, and S. Subramani. 1993. J. Cell Biol. 121:761-774). The PTS1R mRNA is expressed in all human tissues examined. Antibodies to the human PTS1R recognize this protein in human, monkey, rat, and hamster cells. The protein is localized mainly in the cytosol but is also found to be associated with peroxisomes. Part of the peroxisomal PTS1R protein is tightly bound to the peroxisomal membrane. Antibodies to PTS1R inhibit peroxisomal protein-import of PTS1-containing proteins in a permeabilized CHO cell system. In vitro-translated PTS1R protein specifically binds a serine-lysine-leucine-peptide. A PAS8-PTS1R fusion protein complements the P. pastoris pas8 mutant. The PTS1R cDNA also complements the PTS1 protein-import defect in skin fibroblasts from patients--belonging to complementation group two--diagnosed as having neonatal adrenoleukodystrophy or Zellweger syndrome. The PTS1R gene has been localized to a chromosomal location where no other peroxisomal disorder genes are known to map. Our findings represent the only case in which the molecular basis of the protein-import deficiency in human peroxisomal disorders is understood.

Adrenoleukodystrophy↗
PubMed1995

Interaction of alpha-actinin with the cadherin/catenin cell-cell adhesion complex via alpha-catenin.

Cadherins are Ca(2+)-dependent, cell surface glycoproteins involved in cell-cell adhesion. Extracellularly, transmembrane cadherins such as E-, P-, and N-cadherin self-associate, while intracellularly they interact indirectly with the actin-based cytoskeleton. Several intracellular proteins termed catenins, including alpha-catenin, beta-catenin, and plakoglobin, are tightly associated with these cadherins and serve to link them to the cytoskeleton. Here, we present evidence that in fibroblasts alpha-actinin, but not vinculin, colocalizes extensively with the N-cadherin/catenin complex. This is in contrast to epithelial cells where both cytoskeletal proteins colocalize extensively with E-cadherin and catenins. We further show that alpha-actinin, but not vinculin, coimmunoprecipitates specifically with alpha- and beta-catenin from N- and E-cadherin-expressing cells, but only if alpha-catenin is present. Moreover, we show that alpha-actinin coimmunoprecipitates with the N-cadherin/catenin complex in an actin-independent manner. We therefore propose that cadherin/catenin complexes are linked to the actin cytoskeleton via a direct association between alpha-actinin and alpha-catenin.

Actinin↗
PubMed1995

Developmentally regulated mouse gene NK10 encodes a zinc finger repressor protein with differential DNA-binding domains.

Using oligonucleotides complementary to the conserved inter-finger region of a variety of previously described zinc finger-encoding genes, a novel mouse gene was cloned and characterized. The gene is localized on chromosome 8 and comprises five exons. Its corresponding mRNA is developmentally regulated in various tissues and includes an open reading frame encoding a protein of 72,422 daltons. It shares amino-terminal homologies with human KRAB (or FPB) boxes, and contains 13 zinc fingers of the C2-H2 type. The NK10 KRAB domains exhibit repressing activity when tested in GAL4 fusion protein assays. Cloning of putative target sequences revealed that the individual domains differentially contribute to zinc-dependent target DNA binding.

Amino Acid Sequence↗
PubMed1994

Position 713 is critical for catalysis but not iron binding in soybean lipoxygenase 3.

The role of asparagine-713 in iron atom incorporation and catalysis in soybean lipoxygenase 3 was investigated using site-directed mutagenesis. A full-length cDNA for the lipoxygenase isoenzyme was obtained from a library derived from soybeans cv. Provar. Protein with native specific activity at pH 7.4 was obtained from expression in Escherichia coli. Two recent structure reports provided conflicting views about the participation of the side chain of asparagine-694 in the coordination of the iron atom required for catalysis by lipoxygenase 1. Oligonucleotide-directed mutagenesis was employed to modify residue 713 in lipoxygenase 3 which corresponds to asparagine-694 in the sequence of lipoxygenase 1. It was found that for enzyme expressed in bacteria, asparagine was not required for iron incorporation. Histidine, alanine, and serine substitutions for asparagine-713 all produced proteins that contained iron. The histidine mutant had specific activity and catalytic characteristics comparable to the wild-type enzyme. By contrast, the alanine- and serine-substituted lipoxygenases had no detectable catalytic activity. When oxidized by product, the histidine mutant also displayed the characteristic g6 signal of the soybean enzyme in its EPR spectrum. The possibilities that the residue at position 713 acts as a metal ligand, an acid-base catalyst, and a hydrogen bonding group are considered and discussed.

Asparagine↗
PubMed1994

Chromosomal assignment of three novel mouse genes expressed in testicular cells.

The chromosomal positions of three genes that are selectively expressed in mouse testis cells have been identified. These genes include (i) TAZ83, which codes for an early- to mid-pachytene germ cell stage-expressed, cysteine-rich transmembrane protein (cyritestin) with homologies to various snake toxins and guinea pig sperm-egg fusion proteins; (ii) TNZ1, which is expressed in neonatal Leydig cells; and (iii) TAZ4, a testis-specific gene isolated by immunoscreening with antiserum raised against Sertoli cell membranes. Our experimental data, derived from chromosomal in situ hybridizations and RFLP studies of genetic backcrosses, indicate that (i) the TAZ83 (cyritestin) gene maps to chromosome 8, band A2, near the Plat locus; (ii) TNZ1 is located in the proximal region of chromosome 11; and (iii) TAZ4 is located at band D in the distal portion of chromosome 11, near the Hlr1 locus, with a related sequence, TAZ4-rs1, in the proximal part of chromosome 1.

Animals↗
PubMed1994

Jaw muscle function and wishboning of the mandible during mastication in macaques and baboons.

An analysis of in vivo bone strain indicates that the mandibular symphysis of macaques experiences lateral transverse bending or "wishboning" during the power stroke of mastication, and this loading regime results in relatively intense concentrations of stress along the lingual aspect of the symphysis (Hylander 1984, 1985). It has been hypothesized that peak wishboning of the macaque mandible, which probably occurs at the very end of the power stroke, that is, after the initial occurrence of maximum intercuspation, is associated with the late peak activity of the balancing-side deep masseter muscle coupled with the rapid decline in the activity of the balancing-side medial pterygoid and superficial masseter muscles (Hylander et al., 1987). The main purpose of this study is to do a detailed analysis of the electromyographic (EMG) activity of the deep and superficial masseter and medial pterygoid muscles so as to provide a better understanding of the external forces associated with wishboning. This was done by recording and analyzing EMG activity from the superficial masseter, deep masseter, and medial pterygoid muscles in macaques and baboons. EMG activity was recorded from bipolar fine-wire electrodes, and the data were quantified and analyzed using digital techniques. The EMG data clearly support our original observation that the balancing-side posterior deep masseter exhibits peak EMG activity relatively late in the power stroke at a time when activity is rapidly decreasing in the balancing-side medial pterygoid and superficial masseter muscles. Moreover, peak activity of the balancing-side deep masseter occurs at a time when the activity of the working-side deep and superficial masseter and medial pterygoid muscles are also decreasing. Thus, these data are consistent with the hypothesis that wishboning of the mandible is influenced significantly by the late occurrence of force from the balancing-side deep masseter. The EMG data also indicate that residual force from the relaxing superficial masseter may contribute significantly to wishboning. Finally, patterns of wishboning are not fully accounted for by our EMG analysis of the deep and superficial masseter and medial pterygoid muscles. This is probably because the lateral ptergyoids, which can either counter increased wishboning or actually cause reverse wishboning, were not included in our EMG analysis. The EMG data demonstrate that jaw-closing muscle recruitment patterns for macaques and baboons differ from those of humans. Nevertheless, in spite of these differences, it appears, based on previously published EMG data, that the human symphysis may also experience wishboning.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

Mapping of the ACTH, MSH, and neural (MC3 and MC4) melanocortin receptors in the mouse and human.

The melanocortin peptides regulate a wide variety of physiological processes, including pigmentation and glucocorticoid production, and also have several activities in the central and peripheral nervous systems. The melanocortin receptor family includes the melanocyte-stimulating hormone receptor (MSH-R), adrenocorticotropic hormone receptor (ACTH-R), and two neural receptors, MC3-R and MC4-R. In the human these receptors map to 16q24 (MSH-R), 18p11.2 (ACTH-R), 20q13.2 (MC3-R), and 18q22 (MC4-R). The corresponding locations in the mouse are 8, 18, and 2; a variant for mapping MC4-R has not yet been identified. The data reported here also show that the neural MC3 receptor maps close to a disease locus for benign neonatal epilepsy in human and near the E1-2 epilepsy susceptibility locus in the mouse.

Animals↗
PubMed1994

Identification and genetic mapping of 151 dispersed members of 16 ribosomal protein multigene families in the mouse.

More than 150 individual members of 16 ribosomal protein multigene families were identified as DNA restriction fragments and genetically mapped. The ribosomal protein gene-related sequences are widely dispersed throughout the mouse genome. Map positions were determined by analysis of 144 progeny mice from both an interspecific (C57BL/6J x SPRET/Ei)F1 x SPRET/Ei and an intersubspecific (C57BL/6J x CAST/Ei)F1 x C57BL/6J backcross. In addition, 30 members of the multigene families encoding PGK1 ODC, and TPI, including five new loci for ODC and one new locus for TPI, were characterized and mapped. Interspecific backcross linkage data for 29 nonecotropic murine leukemia retroviruses endogenous to C57BL/6J mice are also reported. Transmission ratio distortions and recombination frequencies are compared between the two backcrosses.

Animals↗
PubMed1994

E-cadherin mediates adherens junction organization through protein kinase C.

Cultured human keratinocytes maintained in 30 microM Ca2+ do not form adherens junctions; however, when the extracellular Ca2+ concentration is raised to 1 mM, adherens junctions form very rapidly. The formation of a junction involves the coordinate organization of intracellular and extracellular components. Cadherins have been shown to mediate this coordinate organization. In this report we show that E-cadherin organizes the various junctional components by signalling through protein kinase C.

Cadherins↗
PubMed1993

Characterization of cDNA clones for the 2-methyl branched-chain enoyl-CoA reductase. An enzyme involved in branched-chain fatty acid synthesis in anaerobic mitochondria of the parasitic nematode Ascaris suum.

The 2-methyl branched-chain enoyl-CoA reductase plays a pivotal role in the reversal of beta-oxidation operating in anaerobic mitochondria of the parasitic nematode Ascaris suum. An affinity-purified polyclonal anti-serum against the reductase was used to screen a cDNA library constructed in lambda gt11 with poly(A)+ RNA from adult A. suum muscle. A 1.2-kilobase partial cDNA clone was isolated, subcloned, and sequenced in both directions. Additional sequence at the 5' end of the mRNA was determined by the RACE (rapid amplification of cDNA ends) procedure. Nucleotide sequence analysis of the cDNAs revealed the 22-nucleotide trans-spliced leader sequence characteristic of many nematode mRNAs, an open reading frame of 1236 nucleotides and a 3'-untranslated sequence of 109 nucleotides including a short poly(A) tail 14 nucleotides from a polyadenylation signal (AATAAA). The open reading frame encoded a 396-amino acid sequence (M(r) 43,046) including a 16-amino acid leader peptide. Two-dimensional gel electrophoresis of the purified reductase yielded multiple spots with two distinct but overlapping amino-terminal amino acid sequences. Both sequences overlapped with the sequence predicted from the mRNA, and one of the sequences was identical to the predicted sequence. Comparison of the ascarid sequence with that of mammalian acyl-CoA dehydrogenases revealed a high degree of sequence identity, suggesting that these enzymes may have evolved from a common ancestral gene even though the ascarid enzyme functions as a reductase, not as a dehydrogenase. Immunoblotting of A. suum larval stages and adult tissues with antisera that cross-reacted with each of the spots separated on two-dimensional gels suggested that the reductase was only found in adult muscle. Northern blotting using the partial cDNA revealed a hybridization band of about 1.5 kilobases and also suggested that the enzyme was tissue-specific and developmentally regulated in agreement with the results of the immunoblotting.

Amino Acid Sequence↗