Biomedical subjects

D W Martin

Publications and source records attributed to D W Martin.

At least 145 records · Page 8Linked to original sources

PubMed1976

Increased PRPP synthetase activity in cultured rat hepatoma cells containing mutations in the hypoxanthine-guanine phosphoribosyltransferase gene.

Nine independently derived clones of mutagenized rat hepatoma cells selected for resistance to 6-mercaptopurine (6-MP) or 6-thioguanine (6-ThioG) have been isolated. Each has severely reduced catalytic activity of hypoxanthine-guanine phosphoribosyltransferase (HPRT) and seven of them possess significantly increased activities of phosphoribosylpyrophosphate (PRPP) synthetase. The degrees of elevations of PRPP synthetase activities do not correlate with the degrees of deficiencies of HPRT activities. The cells from one of these clones, 1020/12, posses 40% of the normal HPRT catalytic activity and overproduce purines. We have extensively examined the cells from this clone. Immunotration studies of 1020/12 cells indicate that there is a mutation in the structural gene for HPRT. Although they possess increased specific catalytic activities of the enzyme. PRPP synthetase, the catalytic parameters, heat stability, and isoelectric pH of PRPP synthetase from 1020/12 cells are indistinguishable from those of the enzyme from wild-type cells. The cause of purine overproduction by 1020/12 cells appears to be the elevated PRPP synthetase activity, rather than a PRPP "sparing" effect stemming from reduced HPRT activity. Support for this idea is provided by the observation that the complete loss of HPRT activity in a clone derived from 1020/12 cells does not further enhance the levels of PRPP synthetase or purine overproduction. We propose that the elevated levels of PRPP synthetase activity in these HPRT deficient cells result from a mutational event in the structural gene for HPRT, and that this causes the disruption of a previously undescribed regulatory function of this gene on the expression of the PRPP synthetase gene.

Amidophosphoribosyltransferase↗

PubMed1976

Characterization of a cell culture model for the study of adenosine deaminase- and purine nucleoside phosphorylase-deficient immunologic disease.

The absence of erythrocytic adenosine deaminase (ADA) or purine nucleoside phosphorylase (PNP) has been associated with severe immunodeficiency disease in children. We have developed a cell culture model to study the possible relationships between purine salvage enzymes and immunologic function using an established T cell lymphosarcoma (S49) and a potent inhibitor of ADA, erythro-9(2-hydroxy-3-nonyl) adenine (EHNA). Wild-type S49 cells are killed by dexamethasone or dbc AMP, and adenosine (5 muM) in the presence of an ADA inhibitor (6 muM EHNA) also prevents the growth of and kills these S49 cells. It has been proposed that adenosine is toxic to lymphoid cells by virtue of its ability to increase the intracellular concentrations of cyclic AMP. We examined the sensitivity of three mutants of S49 cells, with distinctive defects in some component of cyclic AMP metabolism or action, to killing by adenosine and EHNA. All three mutants are resistant to killing by isoproterenol or cholera toxin and two are resistant to dbc AMP itself, but all are sensitive to killing by adenosine and EHNA. Similarly, two dexamethasone-resistant S49 mutants are as sensitive to adenosine and EHNA as are the wildtype cells. We have also simulated the purine nucleoside phosphorylase deficiency in S49 cells by adding inosine and adenosine to the growth medium. In the presence of EHNA or inosine, the toxic effects of adenosine can be partially reversed by addition of (10-20 muM) uridine, an observation suggesting that adenosine is toxic as the result of its inducing pyrimidine starvation.

Adenine↗

PubMed1976

Binding of an opiate, levorphanol, to intact neuroblastoma cells in continuous culture.

Mouse neuroblastoma cells in continuous culture, incubated for 1 to 2 days in the presence of 10--6 M levorphanol or morphine, were found to become tolerant to and dependent on those biologically active opiates. Examination of the interaction between levorphanol and the whole neuroblastoma cell suggested that levorphanol was binding to stereospecific opiate receptor sites. This binding was time and temperature dependent, and saturable at concentrations greater than 10--5 M levorphanol. Competition by other opiates for levorphanol sites correlated with their biological activity. This is the first evidence for saturable, stereospecific opiate binding in a homogeneous population of unhybridized cells in continuous culture.

Animals↗

PubMed1976

Evidence for nuclear sites of stereospecific opiate binding in neuroblastoma cells in continuous culture.

A class of stereospecific, saturable receptors for narcotic drugs has been found in a clone of mouse neuroblastoma cells in continuous culture. Cells grown in the presence of 10(-8) M etorphine, a synthetic opiate, had an increased doubling time. The neuroblastoma cell nucleus was found to be the sole site for the high affinity binding of etorphine. It was found that these nuclear receptors (1) became saturated at a concentration of 2 X 10(-9) M etorphine, (2) had a dissociation constant of 5 X 10(-11) M etorphine, (3) were capable of binding etorphine stereospecifically at 37 degrees C but not at 4 degree C, and t4) were protein in nature as evidenced by treatment with proteolytic enzymes. More importantly, the receptor activity appeared to be chromatin-associated.

Animals↗

PubMed1975