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Biomedical subjects

T A Chen

Publications and source records attributed to T A Chen.

29 records · Page 2Linked to original sources
PubMed1987

Rapid and reversible changes in nucleosome structure accompany the activation, repression, and superinduction of murine fibroblast protooncogenes c-fos and c-myc.

A procedure for the isolation of transcriptionally active nucleosomes was used to monitor changes in chromatin structure during the activation, repression, and superinduction of the protooncogenes c-fos and c-myc. Nuclei were isolated from murine fibroblasts at successive times after stimulation of quiescent cell cultures with serum or platelet-derived growth factor. The nucleosomes released by a brief micrococcal nuclease digestion were fractionated by HgII-affinity chromatography to separate the unfolded nucleosomes of transcriptionally active genes (in which the sulfhydryl groups of histone H3 are accessible for binding to HgII) from the compactly beaded nucleosomes of transcriptionally inert DNA sequences (in which the H3 sulfhydryl groups are not accessible). The DNA sequence contents of the HgII-bound and unbound nucleosome fractions were compared by slot-blot hybridizations to 32P-labeled cloned probes for c-fos and c-myc. The binding of the c-fos and c-myc nucleosomes to the HgII column accurately reflected both the timing and the degree of their expression, as determined by run-off transcription assays with the isolated nuclei. The superinduction of c-fos and c-myc expression by an inhibitor of protein synthesis (cycloheximide) was reflected in the persistence of the unfolded, transcriptionally active state of their component nucleosomes. These results provide direct evidence that rapid and reversible changes in nucleosome topography accompany the program of oncogene expression, and they suggest a way to monitor aberrant gene activity during malignant transformation.

Animals↗
PubMed1987

In vitro detection of plant mycoplasmas by immunofluorescent staining with monoclonal antibodies.

Hybridoma techniques have been applied to develop specific monoclonal antibodies (McAbs) against spiroplasmas and mycoplasma-like organisms (MLOs). Advantages of the McAbs have been clearly demonstrated in the detection of aster yellows (AY) MLO by indirect enzyme-linked immunosorbent assay (ELISA) and immunofluorescent staining, compared with conventionally produced polyclonal antibodies. McAbs have also been applied in the in vitro detection and identification of spiroplasmas and AY MLOs on membrane filters using modified direct-count immunofluorescent staining. Organisms were fixed with glutaraldehyde and treated with McAbs and fluorescein isothiocyanate (FITC)-conjugated antimouse immunoglobulin, by incubating and concentrating them in a microcentrifuge tube before collecting them on polycarbonate membrane. This procedure best conserves the helical integrity of spiroplasmas without altering the configuration of surface antigens recognized by McAbs. This method could be valuable in monitoring the fastidious mycoplasmas during purification or cultivation attempts.

Antibodies, Bacterial↗
PubMed1986

Reassembly of c-myc and relaxation of c-fos nucleosomes during differentiation of human leukemic (HL-60) cells.

Human promyelocytic leukemic (HL-60) cells have amplified c-myc protooncogene sequences which lead to an elevated level of c-myc gene expression. Induction of HL-60 cells by phorbol esters to undergo monocytic differentiation results in the suppression of c-myc, but the activation of c-fos gene transcription. Chromatin structures of c-myc and c-fos were compared by measuring their sequences in nucleosome-associated DNA fragments. These nucleosomal particles were released from chromatin by micrococcal nuclease digestion and subsequently analyzed with two dimensional gel electrophoresis. C-myc related sequences were detected in nucleosomal DNA fragments of differentiated cells only, while the c-fos related sequences were found in nucleosomal DNAs of noninduced HL-60 cells. Since the enzyme preferentially digests relaxed DNAs, these results suggest that nucleosomal subunits of c-myc and c-fos chromatin are relaxed during the state of active transcription, and reassembled once their transcription is repressed.

Cell Differentiation↗
PubMed1983

Nutritional requirements of two flower spiroplasmas and honeybee spiroplasma.

A chemically defined medium (CC-494) was used to study the nutritional requirements of three spiroplasmas representing three distinct serogroups: flower spiroplasmas [Spiroplasma floricola and FS (SR-3)] and honeybee spiroplasma [HBS (AS-576)]. Glucose, fructose, and mannose were utilized by all three spiroplasmas. In addition, the honeybee spiroplasma could ferment trehalose, FS (SR-3) could ferment sucrose, and S. floricola could ferment trehalose, sucrose, and raffinose. The three spiroplasmas varied greatly in their requirements of amino acids for growth. S. floricola was the only strain that utilized arginine. HBS (AS-576) required at least one purine and one pyrimidine base (either free base or ribonucleoside) for growth, while both flower spiroplasmas grew with only one base in the medium. Oleic acid, cholesterol, and bovine serum albumin were essential to all three spiroplasmas. Palmitic acid, which was nonessential, promoted growth significantly.

Amino Acids↗
PubMed1982

Spiroplasmas: cultivation in chemically defined medium.

A chemically defined medium, CC-494, supports the cultivation in vitro of several spiroplasmas belonging to three distinct serogroups. Medium CC-494 supports the growth of flower spiroplasmas FS 23-6 and FS SR-3 and of honey bee spiroplasma HBS AS-576. The maximal populations of the two flower spiroplasmas and honey bee spiroplasma are comparable to those cultured in the undefined medium C-3G. The growth rate for all three spiroplasmas is slower in the defined medium.

Culture Media↗
PubMed1978

Effect of osmotic potential, pH, and temperature on the growth of a helical, motile mycoplasma causing corn stunt disease.

Growth characteristics of corn stunt spiroplasma, a helical, motile mycoplasma, were studied over a range of osmolality, pH, and temperature in a simple medium containing 20% (v/v) agamma horse serum, 1.5% (w/v) PPLO broth, and various concentrations of sucrose. The spiroplasma was able to grow in a wide spectrum of osmolalities from 360 to 1120 mosm. Optimal growth was observed in media that contained 0.25-0.35 M sucrose. The organism became longer and thinner in media adjusted to 0.65 M sucrose or more. The spiroplasma lost helicity and motility immediately after transfer to media at pH 5.4 or lower. Optimal pH for growth was 7.2. No growth was observed at pH lower than 5.4 or higher than 8.0. Optimal temperature for growth was 32 degrees C. Very little or no growth was observed at temperatures lower than 15 degrees C or higher than 35 degrees C.

Culture Media↗
PubMedDate not supplied

Substrate utilization in defined media.

Substrate utilization in defined media for two flower spiroplasmas (S. floricola and FS SR-3) and honeybee spiroplasma (HBS AS-576) was investigated. Glucose, fructose, and mannose were utilized by all three spiroplasmas. In addition, HBS (AS-576) could ferment trehalose; FS (SR-3), sucrose; and S. floricola, trehalose, sucrose, and raffinose. The three spiroplasmas varied greatly in growth requirements for amino acids. Only S. floricola utilized arginine. HBS (AS-576) required at least one purine and one pyrimidine base for growth, while both flower spiroplasmas grew with only one base in the medium. Oleic acid, cholesterol, and BSA were essential to all three spiroplasmas. Palmitic acid, which was non-essential, promoted growth significantly.

Amino Acids↗
PubMedDate not supplied

Serological classification of spiroplasmas: current status.

Data concerning serological classification of spiroplasmas are in good agreement, but slightly different numerical designations have been given to existing groups. It is proposed that a standardized system be adopted based on information developed mainly by the IRPCM working team on spiroplasmas. The type species (Spiroplasma citri) should be redefined to include only the agent of citrus stubborn disease (subgroup I-1). Six other subgroups, including three proposed by Bové et al. in this volume (I-5, I-6, and I-7), are members of the Group I complex. Because subgroups I-1, I-2, and I-3 (1) show significant reciprocal differences in DNA-DNA homology and two-dimensional electrophoretic protein profiles, (2) occupy exclusive habitats, (3) are each associated with important diseases, and (4) consist of clusters of very similar or identical strains, it is suggested that Latin binomials could be assigned to subgroups I-2 and I-3. It is proposed that those criteria could serve as general guidelines for consideration of subgroups for species status in the class Mollicutes. The I-4 subgroup is assigned an uncertain status, pending comparisons with the LB-12 (I-5), M55 (I-6), and N525 (I-7) subgroups. To previously described serogroups we add the CN-5 Cotinus beetle spiroplasma (IX), the AES-1 mosquito strain (X), and the MQ-4 Monobia strain (XI).

DNA, Bacterial↗
PubMedDate not supplied

Transcriptional activation of the c-myc oncogene in the T-47D mammary cancer cells by conditioned media from embryonic mouse fibroblasts (BALB/c-3T3). Effect of the antiestrogen ICI 164,384.

The effects of the conditioned medium (CM) of embryonic mouse fibroblast (BALB/c-3T3) cells (clone A31) on the mRNA of the c-myc oncogene of the hormone-dependent T-47D mammary cancer cells were explored. After 6h incubation a significant stimulation of c-myc expression was observed with a 10% v/v concentration of CM, and the effect was even higher using a CM concentration of 30% v/v. It is suggested that this action of CM could be related to the stimulatory effect on proliferation and DNA synthesis induced by the CM, as was recently demonstrated for the T-47D cells. It was also observed that the antiestrogen ICI 164,384 (5 x 10(-6) M) can inhibit the stimulatory action of CM on c-myc expression. It is concluded that the fibroblast mouse embryonic cells contain and secrete factor(s) involved in the mechanism of proliferation of human hormone-dependent mammary cancer cells.

3T3 Cells↗