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N L Anderson

Publications and source records attributed to N L Anderson.

At least 73 records · Page 4Linked to original sources
PubMed1984

An electrophoretic variant of a human fibroblast protein with characteristics of smooth muscle tropomyosin.

A charge variant of a protein (Tm: 3; molecular weight approximately 35,000) that co-migrates with human smooth muscle tropomyosin has been found in whole cell extracts from the fibroblasts of a father and his son. The variant protein co-purifies with Tm: 3, shifts with it to a higher apparent molecular weight on sodium dodecyl sulfate/polyacrylamide gel electrophoresis in the presence of 4 M-urea, is a component of the microfilaments, and has a peptide cleavage pattern identical to that of Tm: 3 and smooth muscle tropomyosin. The results indicate that Tm: 3 and the variant (Tm: 3.1) are smooth muscle tropomyosin, suggesting that normal human fibroblasts synthesize at least two tropomyosins (Tm:3 and the non-muscle tropomyosin Tm:4, molecular weight approximately 30,000) and that they are the products of separate genes.

Amino Acids↗
PubMed1984

Protein-pattern changes and morphological effects due to methionine starvation or treatment with 5-azacytidine of the phorbol-ester-sensitive cell lines HL-60, CCL-119, and U-937.

Methionine starvation causes changes in the protein pattern of HL-60 promyelocytic leukemia cells as observed by two-dimensional electrophoresis. One group of proteins is apparently modified, appearing in new positions. A further series of proteins, including several principal nuclear polypeptides, is substantially diminished. The morphology of a fraction of the cells in the culture changes concomitantly, with condensation and fragmentation of the nucleus and eventual remolding of the cell to a "grape-cluster" appearance. Similar effects are produced by a DNA methylation inhibitor, 5-azacytidine, but not by various other toxic agents tested. A defect in DNA methylation, either by depletion of S-adenosyl-L-methionine (the methyl donor) or by inactivation of the relevant enzyme, may be responsible. The T-lymphoblastoid line CCL-119 and the histiocytic lymphoma line U-937 also show these effects, but most fibroblast, epithelial, and lymphoblastoid lines do not. These changes can be largely prevented in each of the three susceptible lines by prior treatment with the tumor promoter, phorbol myristate acetate (PMA), an agent known to cause differentiation in at least two of the lines. The results thus suggest interesting relationships between methionine metabolism, protein and structural changes in the cell nucleus, and PMA-induced cell differentiation.

Azacitidine↗
PubMed1984

Global approaches to quantitative analysis of gene-expression patterns observed by use of two-dimensional gel electrophoresis.

A major difficulty in the use of two-dimensional protein maps to identify and classify cell types is the problem of acquiring, selecting, and analyzing quantitative data on hundreds of protein spots. Here we use methods of multivariate statistics to analyze the differences among a panel of human cell lines, in some cases involving quantitative data on more than 250 proteins. Principal-component and cluster-analysis techniques show that the lines can be easily distinguished, even by using the subset of proteins present in all cells. A preliminary analysis of the protein changes brought about by phorbol ester-induced differentiation of the line U937 is included.

Cell Differentiation↗
PubMed1984

Two-dimensional electrophoresis used to differentiate the causal agents of American tegumentary leishmaniasis.

American tegumentary leishmaniasis is caused by a diverse group of Leishmania classified within two species complexes: L. mexicana and L. braziliensis. Because distinct disease forms are associated with certain species or subspecies, prognosis requires taxonomic identification of the pathogen. Biological criteria allow identification only to the species level; thus, biochemical and immunological markers are eagerly sought. We have used two-dimensional electrophoresis to examine the relationships among reference strains of New World Leishmania and stocks isolated from Colombian patients. The L. mexicana and L. braziliensis species complexes are shown to be extremely disparate, and the relative affinities of the subspecies reference strains and Colombian Leishmania stocks are documented. The latter displayed a variety of patterns--some clearly identifiable with a particular L. braziliensis subspecies reference strain, others intermediate between those of L.b. panamensis and L.b. guyanensis. Such comparisons are useful both in establishing relatedness and identifying subspecies and variant marker proteins, which may have biological significance.

Animals↗
PubMed1983

Control of macrophage cell differentiation in human promyelocytic HL-60 leukemia cells by 1,25-dihydroxyvitamin D3 and phorbol-12-myristate-13-acetate.

Human promyelocytic leukemia cells (HL-60) were induced to differentiate into macrophage-like cells in a dose (3 X 10(-10) to 10(-7) M) and time (1 to 6 days)-dependent manner by 1,25-dihydroxyvitamin D3 and the tumor promoter, phorbol-12-myristate-13-acetate. Differentiation was determined by an increase in the percentage of morphologically mature cells, in lysozyme and nonspecific esterase activities, and in reactivity with the murine OKM1 monoclonal antibody. Two HL-60 cell variants, designated as R-80 and B-II, were also examined. R-80 cells, which are resistant to induction of cell differentiation by phorbol-12-myristate-13-acetate, also exhibited resistance, although to a lesser degree, to induction of cell differentiation by 1,25-dihydroxyvitamin D3. The resistance to the action of the two compounds is presumably not due to similar binding sites for the two inducers, since 1,25-dihydroxyvitamin D3 was unable to compete for the phorbol diester binding sites as measured by [3H]phorbol-12,13-dibutyrate binding. B-II cells were resistant to induction of cell differentiation by 1,25-dihydroxyvitamin D3, phorbol-12-myristate-13-acetate, retinoic acid, and dimethyl sulfoxide. Two-dimensional electrophoretic analysis of HL-60 cell protein patterns indicated that treatment of the HL-60 cells with 1,25-dihydroxyvitamin D3, phorbol-12-myristate-13-acetate, retinoic acid, and dimethyl sulfoxide caused the cells to express various monocyte-macrophage and granulocyte marker proteins. None of the inducers caused a protein pattern identical to that of peripheral monocytes or granulocytes in the HL-60 cells, but the protein pattern of the HL-60 cells treated with 1,25-dihydroxyvitamin D3 was the closest to that of peripheral blood monocytes. These results indicate that 1,25-dihydroxyvitamin D3 induces in the HL-60 cells a phenotype that resembles, but is not identical to, that of peripheral monocytes-macrophages.

Antibodies, Monoclonal↗
PubMed1983

Analysis of human leukemic cells by use of high-resolution two-dimensional electrophoresis. I: results of a pilot study.

We analyzed mononuclear leukocytes from patients with various human leukemias by high-resolution two-dimensional electrophoresis. Tumor cells of the granulocytic, monocytic, and lymphoid lineages [obtained from chronic granulocytic leukemia in blast transformation, acute monocytic leukemia, and chronic lymphocytic leukemia (CLL), respectively] can be easily recognized by using a series of cell-type marker proteins identified by comparison of fractionated normal cell populations. B and T cell types of CLL could be distinguished, the results correlating well with those obtained by use of monoclonal-antibody staining methods. In two cases representing almost pure B-cells (classical CLL; 0% T, 85% B) and T-cells (cutaneous T-cell leukemia; 77% T, 0% B), 27 of 29 marker proteins showed quantitative B/T differences comparable to those observed in comparisons of normal B-and T-lymphocytes prepared by cell sorting. These results indicate that cells from relatively well-differentiated leukemias show complex patterns of gene expression very similar to those of the corresponding normal cells and strongly support the use of large marker panels in cell-type determination. Less-well-differentiated acute leukemias [such as acute undifferentiated and acute granulocytic (FAB:M1)] appear to yield protein patterns corresponding less closely to recognizable mature cell types, and may show expression of novel proteins related to the state of differentiation.

Adult↗
PubMed1982

Lymphocyte, monocyte, and granulocyte proteins compared by use of two-dimensional electrophoresis.

We compared cellular proteins from normal human blood lymphocytes, monocytes and granulocytes, using high-resolution two-dimensional electrophoresis. The leukocytes were isolated from peripheral blood by centrifugation on density step gradients (yielding a fraction of purified granulocytes and a lymphocyte/monocyte mixture) and monocytes were subsequently separated from lymphocytes by virtue of their adherence to plastic. Wright-stained smears indicated that each of the three resulting fractions was 90 to 95% pure. The cells were labeled with [35S]methionine after various intervals in culture, then solubilized and analyzed by two-dimensional electrophoresis. Although most proteins in each cell type are common to all three, there are nevertheless several specific marker proteins that distinguish one cell type from another. We also examined the appearance of these markers in three lines of cultured cells from humans (GM607, a B-lymphoblastoid line; HL-60, a promyelocytic leukemic line; and 1494, a normal skin fibroblast).

Blood Proteins↗
PubMed1982

Design and implementation of a prototype Human Protein Index.

This paper describes information-handling aspects of the TYCHO I analysis system (Clin, Chem. 27: 1807--1820, 1981), which analyzes two-dimensional electrophoresis gels, matches the individual protein spots with those in a reference pattern, and stores various information--including spot measurements, identifications, treatment profiles, set memberships, and comments--in a computerized database. This and additional information such as amino acid composition and cellular localization is then accessible from an interactive program that includes a pictorial user interface and presents much of the data in graphical form. Use of the TYCHO I system is illustrated by examples drawn from analyses of gel patterns from human leukocytes.

Blood Proteins↗
PubMed1982

A two-dimensional electrophoretic analysis of the heat-shock-induced proteins of human cells.

Using two-dimensional electrophoresis, we have investigated the responses of human cells in culture to heat shock and to various chemical agents producing a similar effect. These treatments result in the induction of increased synthesis of several specific proteins. One (HShock:1, SDS-molecular mass about 65000) is increased by about 350-fold over the amount in untreated cells. Computer analysis of time-course studies indicates, however, that rates of synthesis of various proteins other than the classical "heat shock proteins" are affected, some of these alterations following time courses quite different from the main (HShock) inductions. The heat shock effect is thus much more complicated than previously realized. We purified the HShock:1 protein from heat-shocked human lymphoblastoid cells, and prepared a rabbit antiserum specific for HShock:1 on nitrocellulose two-dimensional gel transfers of total lymphoblastoid cell protein. A survey of mouse tissues shows high concentrations of an HShock:1-like protein in the testis, and human testes also appears to contain substantial (though lower) concentrations. These results are consistent with the hypothesis (derived from the tissue-culture studies) that the heat shock effect is a general response to the need for increased protein catabolism within the cell. Increased concentrations of HShock:1 are also observed in preparations of blood leukocytes collected from patients after surgery, indicating that some types of physiological trauma may induce the heat shock proteins in man. Using the antiHShock:1 antibody in an immunoassay, it will be possible to systematically examine HShock:1 concentrations in plasma and leukocytes, thereby opening up the possibility of a clinical test based for the first time upon an inducible aspect of cellular gene expression.

Animals↗
PubMed1982

Cytoskeletal proteins from human skin fibroblasts, peripheral blood leukocytes, and a lymphoblastoid cell line compared by two-dimensional gel electrophoresis.

Differences in proteins between cells grown as suspension cultures and those grown as attached cultures were studied by comparing the proteins of detergent-resistant cytoskeletons prepared from peripheral blood leukocytes and a lymphoblastoid cell line (GM607) (both grown as suspension cultures) and those of human skin fibroblasts (grown as attached cultures) by two-dimensional gel electrophoresis. The major cytoskeletal proteins of the leukocytes were also present in the protein pattern of GM607 cytoskeletons. In contrast, the fibroblast cytoskeletal protein pattern contained four groups of proteins that differed from the patterns of the leukocytes and GM607. Three groups (Cytoskf:8--10, :14--16, and :17--18) showed qualitative differences, and the fourth group (Cytoskf:11 and :13) showed quantitative differences. In addition, surface labeling of GM607 and human fibroblasts with 125I demonstrated that substantial amounts of vimentin and actin are exposed at the surface of the attached fibroblasts, but there is little evidence of similar exposure at the surface of the suspension-grown GM607. Cytoskf:11 and :13 in fibroblast preparations were also labeled with the 125I. These results demonstrate some differences in cytoskeletal protein composition between different types of cells could be related to their ability or lack of ability to grow as attached cells in tissue culture.

Actins↗
PubMed1981

A variant of human nonmuscle tropomyosin found in fibroblasts by using two-dimensional electrophoresis.

In an analysis of 12 human fibroblast cell lines by two-dimensional electrophoresis, one cell line (1493) was found to contain a major protein variant (Cytosk: 12; for the convention used in spot numbering, see Anderson, N. L. (1981) Proc. Natl. Acad. Sci. U. S. A. 78, 2407-2411) not present in any of the other 11 cell lines. Biochemical characterization of the variant protein included determination of its subcellular location, partial amino acid composition, behavior on sodium dodecyl sulfate (SDS) versus SDS/urea gels, and partial proteolytic digestion patterns. All of these methods showed that Cytosk:12 is related to Cytosk:11. Both proteins are located in the cytoskeleton, contain little cysteine or proline and no detectable tryptophan, shift together to a higher apparent molecular weight when electrophoresed in the presence of SDS and 8 M urea versus SDS alone, and have identical products after partial proteolysis according to Cleveland et al. (Cleveland, D. W., Fischer, S. G., Kirschner, M. W., and Laemmli, U. K. (1977) J. Biol. Chem. 252, 1102-1106). Preparation of nonmuscle tropomyosin resulted in the partial purification of both Cytosk:11 and :12. The data suggest that Cytosk:11 is fibroblast nonmuscle tropomyosin and that Cytosk:12 in cell line 1493 is a charge variant of that protein.

Cell Line↗
PubMed1981

The TYCHO system for computer analysis of two-dimensional gel electrophoresis patterns.

We describe here a computer system for the analysis of high-resolution two-dimensional gel-electrophoresis patterns, with some initial applications. The system (called TYCHO) comprises programs for image acquisition, background subtraction and smoothing, spot detection, gaussian spot modeling, and pattern matching and comparison. It is based on a conventional minicomputer, but makes extensive use of a high-speed array processor in the image-processing and -modeling steps. Used in concert with the ISO-DALT two-dimensional electrophoresis system (Anal. Biochem. 85:331-354, 1978), TYCHO allows quantitative measurement of hundreds of proteins in complex biological samples, and constitutes the initial data-reduction system required for work towards a Human Protein Index.

Blood Proteins↗