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Biomedical subjects

K A Ellem

Publications and source records attributed to K A Ellem.

5 recordsLinked to original sources
PubMed1979

Subfractionation of CsCl-purified H-1 parvovirus on metrizamide gradients.

The different density classes of H-l parvovirus, collected within 30 hr of infection of par-asynchronous cultures, following the standard CsCl purification step, have been shown to be heterogeneous. Rebanding of the denser form (HF, p = 1.46 g/cm3) and the less dense form (LF, p = 1.42 g/cm3) of infectious virus in the nonionic density generating solute, metrizamide, showed that both HF and LF virus bands were heterogeneous in density. The infectivity banded with isotopically labeled virus protein and DNA at 1.32 g/cm3 for both HF and LF virus. Amounts of protein and DNA which varied from preparation to preparation, but which were greater from the HF virus band, were distributed throughout the rest of the gradient, but predominated in a peak at a density of 1.2 g/cm3. The protein in this peak was without hemagglutinating activity but had the molecular weights and proportions of the H-l virion proteins (VPl, VP2', and VPZ). The DNA was of the same size as H-l DNA monomers and its proportion to the protein was similar to that of the infectious peak. The DNA was susceptible to micrococcal nuclease digestion. The nature of this noninfectious viral material thus seemed to be incompletely assembled virus. Radiolabeled H-l virus collected after 72 hr of infection formed a discrete single peak in both CsCl (p = 1.42 g/cm3), and metrizamide gradients (p = 1.32 g/cm3). There was no significant amount of the 1.20 g/cm3 viral protein-DNA complex in these mature preparations.

Centrifugation, Density Gradient↗
PubMed1977

Secondary activities of diverse inhibitors potentiate the response of hamster embryo cultures to a mitotic stimulus.

The reversal of the density dependent inhibition of replication (DDIR) of Syrian hamster embryo cultures by fresh medium containg 30-50% fetal bovine serum was poorly synchronized. There were two waves of DNA synthesis eight to nine hours apart, which by examination of autoradiograms of cultures pulse-labeled with 3H-thymidine, were found to involve 55% and 45% of the cell population. The second wave was found to be due to a subpopulation of epithelioid cells set in the predominantly fibroblastic cell cultures. Pretreatment of the DDIR cultures with hydroxyurea (HU) or arabinosyl cytosine (ara-C), followed by serum stimulation in the absence of the drugs, led to an enhancement of the synchrony. The effect increased with lengthening of the contact with antimetabolites, to a maximum after 20 hours' pre-exposure, and was in part due to the shortening of the GI phase of the epithelioid elements, and in part to increasing the synchrony of the fibroblastic cells. The resulting synchrony involved some 95% of the cells in simultaneous DNA synthesis after a median Gl period of 12 hours. The effect had no relationship to the role of HU and ara-C as specific inhibitors of DNA synthesis since the cultures were mitotically quiescent, and a similar enhanced response could be induced in DDIR cultures by prestimulation exposure lasting only two hours to cycloheximide (cyx), an inhibitor of protein synthesis. Pre-exposure of DDIR cultures to actinomycin D did not potentiate the cell response. A survey of the known secondary inhibitions caused by these three antimetabolites suggests that they all may cause deficiencies in the glycolipid or glycoprotein moieties of the cell surface. These observation provide a use ful, simple means of improving synchrony in these systems and may prove to be a useful probe for investigating the role of the cell surface in regulating cell replication.

Animals↗
PubMed1975

Nutritional effects on precursor uptake and compartmentalization of intracellular pools in relation to RNA synthesis.

The effects of nutritional variables on the processing of exogenous precursors into RNA was examined. General nutritional deprivation, or asparagine depletion, led to significant changes in the absolute pool sizes, especially of ATP, UTP and CTP. Fluctuations were found depending on the elapsed time after the nutritional perturbations occurred, and the cell density of the cultures. Depletion of the medium by 28 h of growth, or 1 h of guinea pig asparaginase action, led to considerable inhibition of the conversion of exogenous uridine to CTP by the cells. A series of experiments indicated that in 6C3HED lymphoma cells the uridine nucleotide pool which provided the immediate precursors to RNA (denoted UTP-NA) behaves as a small compartment in rapid equilibrium with exogenously supplied nucleosides. The resemblance to the compartmentation model described by Plagemann (Plagemann, P.G.W. (1972) J. Cell Biol. 52, 131-146 and (1971) J. Cell. Physiol. 77, 241-258) for rat hepatoma cells was close. The UTP-NA pool of the 6C3HED cells constitutes no more than 5% of the cellular UTP pool and is relatively slow in equilibrating with the general cell pool. Correction of the rates of incorporation of isotope into RNA by using some function of the whole cell UTP specific activity to normalize the pool effects, was shown to be invalid.

Adenosine Triphosphate↗