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Biomedical subjects

J Hu

Publications and source records attributed to J Hu.

At least 613 records · Page 34Linked to original sources
PubMed1989

Binding and transcription of relaxed DNA templates by fractions of maize chloroplast extracts.

Preparations of partially purified chloroplast DNA-dependent RNA polymerase from maize and some other plants transcribe cloned chloroplast genes preferentially and much more actively from appropriately negatively supercoiled templates than from relaxed templates. We have found that the polymerase in such fractions does not bind to promoter regions of the maize chloroplast genes psbA and rbcL on small linear DNA fragments but that some protein(s) in unfractionated chloroplast extracts does bind. DEAE chromatography of the extracts has permitted the separation of a DNA-binding fraction from the bulk of the RNA polymerase activity. The binding fraction contains plastid RNA polymerase activity that is relatively independent of template topology.

Amino Acid Sequence↗
PubMed1988

Tobacco mosaic virus infection stimulates the phosphorylation of a plant protein associated with double-stranded RNA-dependent protein kinase activity.

The influence of tobacco mosaic virus (TMV) infection on nucleotide binding and phosphorylation of an Mr 68,000 host-encoded protein (p68) was examined. The phosphorylation of p68 in homogenates from TMV-infected tissues was 4-fold greater than in homogenates from mock inoculated tissues. Phosphorylation of p68 in extracts from mock inoculated tissues was enhanced by the addition of double-stranded (ds) RNA. Nucleotide photoaffinity labeling experiments indicate that p68 contains an ATP binding site with characteristics consistent with protein kinase activity. Antiserum raised against a dsRNA-dependent protein kinase activity. Antiserum raised against a dsRNA-dependent protein kinase from interferon-treated human cells immunoprecipitated p68 from extracts of TMV-infected tissue, and p68-containing immunocomplexes catalyzed the phosphorylation of endogenous p68. These data suggest that p68 may be an autophosphorylating, dsRNA-dependent protein kinase involved in viral pathogenesis. Based upon analogous functions demonstrated for dsRNA-dependent protein kinases in mammalian systems, p68 may have a role in the regulation of protein synthesis and viral replication in infected cells.

Electrophoresis, Polyacrylamide Gel↗
PubMed1988

Yeast activators stimulate plant gene expression.

GAL4 is a transcriptional activator found in yeast. Two distinct functions of the protein are required for its activity: one directs sequence-specific DNA binding, and another interacts with some other component of the transcriptional machinery, for example, RNA polymerase II or a TATA-binding protein. Two short regions of GAL4 function as 'activating sequences' when attached to the DNA-binding portion of GAL4 and these regions can be replaced by a large number of peptides encoded by Escherichia coli genomic DNA fragments or by a synthetic peptide designed to form an amphiphilic alpha-helix. All of these activating sequences, like that found in another yeast activator, GCN4 bear an excess negative charge. GAL4 and its derivatives that are active in yeast stimulate transcription in mammalian cells when GAL4 binding sites are introduced upstream of a mammalian gene; similarly, GAL4 activates transcription in Drosophila cells. Here we show that GAL4 derivatives stimulate gene expression in plant cells.

Acetyltransferases↗
PubMed1988

Viroid-induced phosphorylation of a host protein related to a dsRNA-dependent protein kinase.

Viroids are very small, unencapsidated RNAs that replicate and induce severe disease in plants without encoding for any proteins. The mechanisms by which the viroid RNA regulates these events and interacts with host factors are unknown. An Mr 68,000 host-encoded protein has been identified that is differentially phosphorylated in extracts from viroid-infected and mock-inoculated tissues. This phosphoprotein is immunologically related to a double-stranded (ds) RNA-dependent protein kinase from virus-infected, interferon-treated human cells. Further, nucleotide photoaffinity labeling indicates that the protein has an ATP binding site. This protein is similar to dsRNA-dependent protein kinases implicated in mammalian systems in the regulation of protein synthesis and virus replication.

Molecular Weight↗
PubMed1988

Activation signals via CD2 molecule and interleukin 2 receptor act in synergy for helper function induction.

The membrane CD2 molecule appears to play an important role in T cell activation. Indeed, T cell stimulation by some combinations of anti-CD2 monoclonal antibodies (mAb) can result in antigen-independent expression of helper function as assessed by proliferation and lymphokine secretion. We report here that T cell stimulation by a combination of two anti-CD2 mAb recognizing GT2 and T11(1) epitopes, respectively, cannot alone induce T helper clones to proliferate when preincubated in culture medium devoid of exogenous interleukin 2 (IL2). The concerted action of both anti-GT2 + T11(1) mAb and exogenous recombinant IL2 is required to induce cloned helper T cells to produce IL 2 and interferon-gamma to significantly increase IL2 receptors (IL2R) and finally to divide by an autocrine mechanism, whereas each signal alone has no effect. This therefore suggests that, under some conditions of CD2 stimulation, two minimal signals may be delivered through CD2 and IL2R and act synergistically to achieve a complete expression of T helper cell functions. Moreover, analysis of phosphatidylinositol and phosphatidic acid metabolic changes mediated by each signal separately or together suggests that, in this model, IL2 increases the phosphoinositide turnover induced by anti-CD2 antibodies up to a level required for helper function acquisition.

Antigens, Differentiation↗
PubMed1988

Isolation and structure of corticostatin peptides from rabbit fetal and adult lung.

A 34-amino acid peptide and three other structurally related peptides were isolated from rabbit fetal and adult lung. These cationic arginine- and cysteine-rich peptides inhibit corticotropin (ACTH)-stimulated rat adrenal cell corticosterone production. The peptide was called corticostatin (CSI). CSI was purified by reverse-phase HPLC and was shown to be homogenous from its amino acid analysis. Its sequence was determined on a gas-phase sequenator. The structure of CSI is Gly-Ile-Cys-Ala-Cys-Arg-Arg-Arg-Phe-Cys-Pro-Asn-Ser-Glu-Arg-Phe-Ser-Gly- Tyr-Cys - Arg-Val-Asn-Gly-Ala-Arg-Tyr-Val-Arg-Cys-Cys-Ser-Arg-Arg. CSI was found to markedly inhibit ACTH-stimulated corticosterone production by rat adrenal cells in vitro but did not affect basal levels. CSI did not affect the stimulation of aldosterone synthesis by angiotensin II in rat zona glomerulosa cells but it did suppress ACTH-stimulated aldosterone synthesis in whole adrenal cells, demonstrating that CSI is a specific inhibitor of ACTH-stimulated corticosteroid synthesis. The minimum effective concentration of CSI inhibiting ACTH-stimulated (33 pM) corticosterone production was 5 nM (20 ng/ml), the ED50 (50% effective dose) was 25 nM and steroidogenesis was completely inhibited at concentrations greater than 500 nM (2 micrograms/ml).

Adrenal Glands↗
PubMed1988

The effects of radiotherapy on the immune system of patients with nasopharyngeal carcinoma.

We report on 123 patients with nasopharyngeal carcinoma whose immune status was measured at the time of diagnosis, the day radiotherapy was completed, and then 2-3 months and 6-8 months after completion of radiotherapy. Immunological tests performed included the lymphocyte transformation test, the erythrocyte-rosette formation test (ERFT), the 29 degrees C erythrocyte-rosette formation test (29 degrees C ERFT), lymphocyte counts (lymphocytes/mm3 and percentage of lymphocytes), levels of serum immunoglobulins (IgG, IgA, IgM), complement (C3) and circulating immune complexes (CIC), the antinuclear antibody test and a skin test using phytohaemagglutinin (PHA). There were statistically significant differences in all tests (except C3) between patients and normal controls. Marked differences were seen in the lymphocyte count, ERFT, and 29 degrees C ERFT after radiotherapy (p less than 0.01). The diameters of induration of the PHA skin tests were less than those before radiotherapy (p less than 0.01). There were higher incidences of recurrence and metastases in the patients with high levels of CIC and low numbers of lymphocytes in the peripheral blood after radiotherapy. Cellular immunity remained at a low level 8 months after radiotherapy.

Antibodies, Antinuclear↗
PubMed1987

Interleukin 2 up-regulates its own production.

It has been previously reported that a combination pair of anti-CD2 monoclonal antibodies (mAb) T11(2)+T11(3) induces a strong proliferation of T cells, which does not require the involvement of accessory cells and exogenous interleukin 2 (IL-2). More recently, we have shown that the requirement for optimal T cell proliferation depends on the combination pairs of anti-CD2 mAb used. Among them, anti-GT2+T11(1) mAb do not allow optimal proliferation of TA4 helper cloned T cells due, at least in part, to a low level of IL-2 production. This observation offered us the opportunity to study the effect of IL-2 on its own production. We show here that stimulation of cloned TA4 cells with anti-GT2+T11(1) mAb induces only a marginal level of IL-2 production. By contrast, significantly higher levels of IL-2 activity are detected in the culture supernatant of TA4 cells preincubated with recombinant IL-2 (rIL-2) before stimulation with anti-GT2+T11(1) mAb. This effect is dose-dependent over a wide range (5 to 50 IU/ml) of rIL-2 concentrations added during preincubation time. In addition, it is not due to carryover of rIL-2 bound during the preincubation time, or to lesser IL-2 consumption by these cells, or to increasing numbers of IL-2-producing cells induced by exogenous IL-2. Moreover, the observation was confirmed with IL-2 mRNA. Although neither rIL-2 nor anti-GT2+T11(1) mAb alone could induce a significant production of IL-2, rIL-2 appears to up-regulate its own production when the TA4 cells are activated by the anti-CD2 mAb-mediated second signal.

Antibodies, Monoclonal↗
PubMed1987

Regulation of helper T cell clone proliferation via the CD2 molecule.

We have investigated the requirements for CD2-induced proliferation of a CD4+, CD8-, CD3+, CD2+ antigen-specific, class II-restricted proliferating cloned cell line. A combination pair of two monoclonal antibodies (MoAb) recognizing, respectively, TII1 and D66 epitopes on the CD2 molecule was used as a stimulus. The regulatory function of accessory cells and various interleukins in this proliferation was determined. The results show that although this clone was able to proliferate in the absence of accessory cells (AC) or interleukin 1 (IL-1) when stimulated by these MoAb, AC constantly enhanced the response to these MoAb. AC acted by increasing high-affinity IL-2 receptor expression. On the contrary they did not play any role in IL-2 production. This regulation of IL-2 receptor expression by AC was specific of adherent cells, did not involve Fc receptors, was impaired when AC were metabolically inactivated and did not require T cell-AC interaction via LFA1, CD4, or HLA molecules. The AC function was not abrogated by anti-IL-1 antibodies and could not be replaced by exogenous IL-1. These results were compared to previously described AC effects on resting T-cell proliferation when stimulated with the same pair of anti-CD2 MoAb. Clear differences in activation requirements in resting and activated T cells via CD2 molecules were found.

Antibodies, Monoclonal↗
PubMed1987

Delayed mutation in Chinese hamster cells.

The possibility was examined that mutational events can be delayed for more than one or two cell divisions following treatment of Chinese hamster cells with the DNA alkylating agent ethyl methane sulfonate. If mutations in mammalian cells are delayed, the proportion of mutant cells in colonies grown from single mutagen-treated cells will reflect the cell division at which the mutation is genetically fixed, i.e., a first division mutation yields a 1/2 mutant colony, a fifth division mutation produces a 1/32 mutant colony, etc. In the present study, replating of cells from single colonies grown for six to seven days after mutagen treatment resulted in the discrete ratios of glucose-6-phosphate dehydrogenase (G6PD)-deficient mutant to wild-type colonies expected for a delayed mutational process which produces mutations over at least 8-10 cell generations. Further, when cells from 7- to 10-day colonies, grown from ethyl methane sulfonate (EMS)-treated cells were replated into selective medium containing 6-thioguanine (6TG), the number of 6TG-resistant colonies obtained per flask was distributed over a very wide range, consistent with a mutational delay process. These results could not be explained by differences in the number of cells per colony or plating efficiency in selective medium. Assuming that the relative number of 6TG-resistant colonies per flask reflects the time of mutation, EMS treatment produced two groups of mutational events: one which occurred within the first five cell generations and another uniformly distributed over at least the next eight to nine divisions. These results support the conclusion that EMS induces mutants for at least 10-14 cell generations after treatment and raise the possibility that current methods to assess the mutagenic potential of an agent might lead to significant underestimation. The role of delayed mutation in the phenomenon of "mutation expression time" is also discussed.

Animals↗
PubMed1987

Normal DNA ligase activity in a gamma-ray-sensitive Chinese hamster mutant.

A Chinese hamster cell mutant (XR-1) was previously described that is extremely deficient in the repair of double-strand DNA breaks produced by gamma-irradiation during the sensitive G1--early-S period and somewhat deficient in repair of gamma-ray-induced single-strand DNA breaks. To determine whether a deficiency in DNA ligase activity might underlie the biochemical defect, protein extracts from mutant and parental cells were examined for their ability to ligate single- and double-strand breaks in DNA. The kinetics of ligation of single 5'-phosphate-3'-hydroxyl breaks in double-stranded DNA were the same in protein extracts from both cells. After separation of protein extracts by gel-filtration chromatography, the percentage of activity in the large and small molecular forms of DNA ligase was also similar in the two cells. Finally, protein extracts prepared from exponentially growing or G1-synchronized mutant and parental cells were equal in their ability to ligate blunt-end DNA substrates. These data suggest that a deficiency in DNA ligase is not the cause of the repair defect in the XR-1 mutant cell.

Animals↗
PubMed1986

Taxonomy of cucurbit-infecting tobamoviruses as determined by serological and molecular hybridization analyses.

A number of cucurbit-infecting tobamoviruses have been reported in the past but there is confusion about their identity and relationships. Cucumber viruses 3 (CV3) and 4 (CV4) were originally described in the United Kingdom whereas the watermelon (W) and cucumber (C) 'isolates' of cucumber green mottle mosaic virus (CGMMV) came from Japan. The results of serological studies and RNA-cDNA analyses have shown that CV3, CV4 and CGMMV-W are very closely related, whereas CGMMV-C is quite different. It is concluded that there are two distinct tobamoviruses that infect cucurbits; they are only very remotely related to each other and to a number of other tobamoviruses. It is suggested that the name CGMMV be retained to include the isolates or strains CV3, CV4 and CGMMV-W, and that a new name, kyuri green mottle mosaic virus (KGMMV), be given to CGMMV-C.

Antigens, Viral↗
PubMed1985

Cell cycle-dependent repair of double-strand DNA breaks in a gamma-ray-sensitive Chinese hamster cell.

A Chinese hamster cell mutant has been isolated which is extremely sensitive to killing by gamma-irradiation in the G1 and early S phases of the cell cycle (LD50 of 20 vs. 250 rads for parent), but which has nearly normal resistance in late S. The mutant cell is able to repair single-stranded DNA breaks introduced by gamma-radiation. However, in comparison to its parental cell, the mutant is deficient in the repair of double-stranded DNA breaks produced by gamma-irradiation during the sensitive G1-early S period, while in the resistant late S period, the repair is nearly the same for both cell types. This correlation between gamma-ray sensitivity and repair strongly suggests that an inability to repair double-strand DNA breaks in G1 is the basis for the hypersensitivity of the mutant to killing by gamma-rays in this phase of the cell cycle. It also provides direct evidence in mammalian cells that the ability to repair double-strand DNA breaks induced by ionizing radiation is an important biochemical function in cell survival and supports the hypothesis that unrepaired double-strand breaks are a major lethal lesion in mammalian cells. A plausible explanation for the appearance of the cell cycle phenotype of the mutant is that in normal cells there are at least two pathways for the repair of double-strand breaks, one of which functions primarily in late S phase, and the other, either throughout the cell cycle or only in the G1 and early S phases.

Animals↗