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Biomedical subjects

R E Palmer

Publications and source records attributed to R E Palmer.

At least 19 recordsLinked to original sources
PubMed1992

Identification of actin and HSP 70 as cyclosporin A binding proteins by photoaffinity labeling and fluorescence displacement assays.

A novel family of cyclosporin A (CsA) binding proteins was identified by using the biologically active, radioiodinated photoaffinity probe [D-Lys-N epsilon-(4-azido-3-[125I]iodophenyl)propionyl)]8-CsA. In addition to cyclophilin, proteins with molecular masses of 43 kDa and approximately 50-55 kDa were labeled in Jurkat extracts and bovine calf thymus. Sequence analysis of the 43-kDa protein purified from calf thymus and subsequent Western analysis of CsA affinity-purified material from Jurkat extracts identified the 43-kDa component as actin. [D-Lys-N epsilon-(5-dimethylamino-1-naphthalenesulfonyl)]8-CsA, a fluorescent analogue of CsA, was prepared and used to measure the binding constants of cyclosporin derivatives to actin by means of a new fluorescence displacement assay. [D-Lys-N epsilon-(5-dimethylamino-1-naphthalenesulfonyl)]8-CsA and [N delta-t-butoxycarbonyl diaminobutyryl)]8-CsA bind to bovine actin at physiologically relevant concentrations, with dissociation constants of 60 +/- 33 and 570 +/- 380 nM, respectively. Because the ATPase fragment of heat shock cognate 70 (HSC 70) is structurally related to actin, the yeast homologue SSA1 was tested and found to be radiolabeled by the cyclosporin A photoaffinity reagent. The binding constant for [D-Lys-N epsilon-(5-dimethylamino-1-naphthalenesulfonyl)]8-CsA to SSA1 was determined and is 53 +/- 48 nM. These results indicate that actin and the 70-kDa heat shock protein family contain a structurally related domain for binding of cyclosporin A-related peptides.

Actins↗
PubMed1992

Role of astral microtubules and actin in spindle orientation and migration in the budding yeast, Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae, before the onset of anaphase, the spindle apparatus is always positioned with one spindle pole at, or through, the neck between the mother cell and the growing bud. This spindle orientation enables proper chromosome segregation to occur during anaphase, allowing one replicated genome to be segregated into the bud and the other to remain in the mother cell. In this study, we synchronized a population of cells before the onset of anaphase such that > 90% of the cells in the population had spindles with the correct orientation, and then disrupted specific cytoskeletal elements using temperature-sensitive mutations. Disruption of either the astral microtubules or actin function resulted in improper spindle orientation in approximately 40-50% of the cells. When cells with disrupted astral microtubules or actin function entered into anaphase, there was a 100-200-fold increase in the frequency of binucleated cell bodies. Thus, the maintenance of proper spindle orientation by these cytoskeletal elements was essential for proper chromosome segregation. These data are consistent with the model that proper spindle orientation is maintained by directly or indirectly tethering the astral microtubules to the actin cytoskeleton. After nuclear migration, but before anaphase, bulk chromosome movement occurs within the nucleus apparently because the chromosomes are attached to a mobile spindle. The frequency and magnitude of bulk chromosome movement is greatly diminished by disruption of the astral microtubules but not by disruption of the nonkinetochore spindle microtubules. These results suggest that astral microtubules are not only important for spindle orientation before anaphase, but they also mediate force on the spindle, generating spindle displacement and in turn chromosome movement. Potential roles for this force in spindle assembly and orientation are discussed.

Actins↗
PubMed1991

Skeletal muscle relaxation with diazo-2: the effect of altered pH.

In a fatigued muscle fibre, the concentrations of ADP, Pi and H+ are all increased and relaxation is slowed. We have used the technique of laser flash photolysis of the caged calcium-chelator, diazo-2, to investigate the direct effect of changes in pH (pH 6.5, 7.0, 7.5) upon tension during relaxation of single chemically skinned fibres, when the effects of the sarcoplasmic reticulum are absent. The relaxation transients were closely fitted with 2 exponentials, a fast (42.3 +/- 1.4; pH 7.0) and a slow process (12.0 +/- 0.7; pH 7.0). The fast phase of relaxation was pH sensitive; lowering pH leading to a slowing of the rate of force decline and raising pH leading to an increase of the rate. The rate of the slow phase was unaltered by changing pH over the range investigated. Thus the slowing of relaxation in fatigued muscle may be due, in part, to the direct action of protons on the myofilaments independent of any effects upon the sarcoplasmic reticulum.

Animals↗
PubMed1990

Striated scallop muscle relaxation: fast force transients produced by photolysis of Diazo-2.

Relaxation of the myosin regulated striated adductor muscles of Pecten maximus was initiated by the photolysis of the caged Ca2+ chelator, Diazo-2. The fibres relaxed to approximately 30% of the maximum tension with a mean half-time of 17.9 +/- 1.6 ms (n = 7, temp 12 degrees C), much faster than the rates observed in intact muscle at the same temperature. This indicates that in the intact adductor muscle the slower relaxation rate is determined by the speed of Ca2+ removal from the sarcoplasm. The faster rate of relaxation of scallop muscle in vitro, compared with frog skeletal muscle may reflect different mechanisms of regulation of the crossbridge cycle.

Animals↗
PubMed1990

Mitotic transmission of artificial chromosomes in cdc mutants of the yeast, Saccharomyces cerevisiae.

In the yeast, Saccharomyces cerevisiae, cell division cycle (CDC) genes have been identified whose products are required for the execution of different steps in the cell cycle. In this study, the fidelity of transmission of a 14-kb circular minichromosome and a 155-kb linear chromosome fragment was examined in cell divisions where specific CDC products were temporarily inactivated with either inhibitors, or temperature sensitive mutations in the appropriate CDC gene. All of the cdc mutants previously shown to induce loss of endogenous linear chromosomes also induced loss of a circular minichromosome and a large linear chromosome fragment in our study (either 1:0 or 2:0 loss events). Therefore, the efficient transmission of these artificial chromosomes depends upon the same trans factors that are required for the efficient transmission of endogenous chromosomes. In a subset of cdc mutants (cdc6, cdc7 and cdc16), the rate of minichromosome loss was significantly greater than the rate of loss of the linear chromosome fragment, suggesting that a structural feature of the minichromosome (nucleotide content, length or topology) makes the minichromosome hypersensitive to the level of function of these CDC gene products. In another subset of cdc mutants (cdc7 and cdc17), the relative rate of 1:0 events to 2:0 events differed for the minichromosome and chromosome fragment, suggesting that the type of chromosome loss event observed in these mutants was dependent upon chromosome structure. Finally, we show that 2:0 events for the minichromosome can occur by both a RAD52 dependent and RAD52 independent mechanism. These results are discussed in the context of the molecular functions of the CDC products.

Cell Division↗
PubMed1989

The dynamics of chromosome movement in the budding yeast Saccharomyces cerevisiae.

Nuclear DNA movement in the yeast, Saccharomyces cerevisiae, was analyzed in live cells using digital imaging microscopy and corroborated by the analysis of nuclear DNA position in fixed cells. During anaphase, the replicated nuclear genomes initially separated at a rate of 1 micron/min. As the genomes separated, the rate of movement became discontinuous. In addition, the axis defined by the segregating genomes rotated relative to the cell surface. The similarity between these results and those previously obtained in higher eukaryotes suggest that the mechanism of anaphase movement may be highly conserved. Before chromosome separation, novel nuclear DNA movements were observed in cdc13, cdc16, and cdc23 cells but not in wild-type or cdc20 cells. These novel nuclear DNA movements correlated with variability in spindle position and length in cdc16 cells. Models for the mechanism of these movements and their induction by certain cdc mutants are discussed.

Cell Cycle↗
PubMed1988

Serum hormone levels associated with spawning activity in the mummichog, Fundulus heteroclitus.

Daily collections of the mummichog, Fundulus heteroclitus, from field populations during the first 45 days of the breeding season revealed a semilunar cycle in the sperm index. Peaks in the sperm index were preceded by 6 days with peaks in the serum testosterone concentration. Bihourly sampling of field populations during a 72-hr period at the new moon showed both diel cycling and an upward trend in serum progesterone levels in female mummichogs. Male mummichogs had 12-hr cycles in serum 17 beta-hydroxy-4-androstene-3,11-dione levels, the peaks of which preceded high tide by 4 hr. The physiological significance of these cycles are discussed.

Androgens↗
PubMed1985

Skull base surgery for glomus jugulare tumors.

Thirty-six patients with glomus jugulare tumors have been managed over a 13-year period using various combinations of skull base surgery and irradiation therapy. The data resulting from this study are presented; the techniques of diagnosis and treatment are reviewed. We conclude that irradiation therapy alone is a satisfactory form of treatment for elderly and poor-risk patients; preoperative x-ray therapy followed by skull base surgery is an effective treatment for younger patients.

Adult↗
PubMed1977

Combined approach surgery for removal of glomus jugulare tumors.

The ideal surgical treatment for glomus jugulare tumors is total removal. Efforts have been made periodically to accomplish this since shortly after this tumor was first identified in the early 1940's. This paper describes a method of removal using a combined approach through the neck and temporal bone, preceded by a course of preoperative irradiation therapy. The early results that have been obtained using this procedure in 10 patients are reported.

Adult↗
PubMed1977

Development of methods for the quantitative in vitro analysis of androgen-dependent and autonomous Shionogi carcinoma 115 cells.

Androgen-dependent and androgen-independent (autonomous), cloned, cultured cell lines of the androgen-dependent mouse mammary adenocarcinoma, Shionogi carcinoma 115, have been established. Growth of the dependent cells requires the presence of androgen, provided they are growth in suspension culture in medium containing dextran-charcoal-treated fetal calf serum. The growth rate of autonomous cells in the presence or absence of DHT is similar to that of dependent cells grown in its presence. An agar culture method has been developed that enables the proportion of dependent and autonomous cells in mixed populations to be determined. Autonomous cells appear in dependent clones, and their frequency increases with increasing time of subculture. Dependent cells form tumors preferentially in male animals and dependent cell cytosols contain significant amounts (approximately 300 femtomoles per mg protein) of a specific androgen-binding macromolecule. Autonomous cells formed tumors equally well in both male and female mice, and autonomous cell cytols contain very low levels (less than or equal to 7 femtomoles per mg protein) of the specific androgen-binding macromolecule(s). These studies delineate a system which can be used to investigate the mechanism of steroid hormone-dependent and autonomous tumor growth, and the transitions between the hormone-dependent and autonomous states.

Animals↗