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Dive into the research topics where Lorraine C. Pfefferkorn is active.

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Experimental Parasitology | 1980

Toxoplasma gondii: genetic recombination between drug resistant mutants.

Lorraine C. Pfefferkorn; E.R. Pfefferkorn

Abstract Mutants resistant to adenine arabinoside (ara-A) or to 5-fluorodeoxyuridine (FUDR) were isolated from a newly isolated oocyst producing strain of Toxoplasma gondii . The selection and characterization of these mutants were carried out in human fibroblast cultures. The ara-A-resistant mutant lacked the enzyme adenosine kinase. The biochemical basis of FUDR resistance remains unknown. Both mutants were used to infect mice to produce brain cysts that contained bradyzoites. Mouse brains that contained cysts were fed to kittens to complete the sexual cycle of T. gondii . Those kittens fed cysts of only one drug-resistant mutant excreted oocysts that yielded no detectable recombinant doubly resistant parasites that could make plaques in the presence of both ara-A and FUDR. Kittens fed a mixture of cysts that contained both mutants excreted oocysts that contained approximately 12% doubly resistant parasites. The reciprocal recombinant, sensitive to both drugs, was also isolated. The doubly resistant recombinant was totally deficient in adenosine kinase activity. This pattern of inheritance is consistant only with a haploid genome for all stages of T. gondii except the zygote formed by fusion of gametes and the unsporulated oocyst. Two FUDR-resistant mutants were also defective in the production of oocysts. These mutants failed to recombine with an ara-A-resistant mutant of proven fertility and thus their inability to make oocysts must result from a defect in the production of both microgametes and macrogametes.


Experimental Parasitology | 1977

Toxoplasma gondii: characterization of a mutant resistant to 5-fluorodeoxyuridine.

E.R. Pfefferkorn; Lorraine C. Pfefferkorn

Abstract An analog of deoxyuridine, 5-fluorodeoxyuridine (FUDR), inhibited the growth of intracellular wild-type Toxoplasma gondii in human fibroblast cultures. FUDR inhibited both RNA and DNA synthesis by the parasite. A single-step FUDR-resistant mutant of T. gondii was isolated in human fibroblast cultures. This mutant was about 100-fold more resistant to FUDR than was the wild-type parasite. To understand the resistance to FUDR we traced the metabolism of deoxyuridine in the host cell and in the parasite. In the host cell, deoxyuridine was specifically incorporated into DNA. In the wild-type parasite deoxyuridine proved to be a general precursor for both RNA and DNA. This pattern of incorporation in T. gondii can be explained if deoxyuridine is converted to uracil, then uridine, and finally uridylic acid in the parasite. The FUDR-resistant but not the wild-type T. gondii was also resistant to fluorouracil and fluorouridine. Autoradiographic analysis showed that intracellular wild-type T. gondii was labeled by deoxyuridine, uracil, or uridine while the FUDR-resistant mutant failed to incorporate any of these precursors. The single genetic defect in salvage or biosynthetic enzymes that is consistent with this pattern of incorporation and resistance is a lack of uridine kinase. However, a general defect in the pyrimidine permeability of the mutant parasite cannot be excluded.


Experimental Parasitology | 1977

Toxoplasma gondii: Specific labeling of nucleic acids of intracellular parasites in Lesch-Nyhan cells

E.R. Pfefferkorn; Lorraine C. Pfefferkorn

Abstract Lesch-Nyhan cells, which are incapable of incorporating hypoxanthine and guanine, supported the growth of Toxoplasma gondii. Massive infection of these cultured mutant human fibroblast cells resulted in a marked increase in the incorporation of radioactive hypoxanthine or guanine supplied in the medium. Autoradiography established that these radioactive purines supplied to infected Lesch-Nyhan cells were incorporated specifically into the intracellular parasites. No such specificity was seen in infected wild-type human fibroblast cells in which the nucleic acids of both the parasite and the host cell were labeled. Purine nucleotides within the intracellular parasite were not available for the synthesis of host nucleic acids. Toxoplasma gondii was less active than wild-type human fibroblast cells in the interconversion of purine nucleotides.


Molecular Immunology | 1990

Functional comparison of the inductions of NADPH oxidase activity and Fcγ RI in IFNγ-treated U937 cells☆

Lorraine C. Pfefferkorn; Paul M. Guyre; Michael W. Fanger

Abstract The capacity to generate Superoxide anion (O − 2 ) can be induced in U937 cells by various agents known to cause myeloid cell differentiation. Other reported differentiation events include diminished cell proliferation and the induction by gamma-interferon (IFNγ) of Fc receptors for immunoglobulin G1 (FcγRI). In this study, we differentiated U937 cells and high FeγRI-expression mutants of U937 cells by treating them with IFNγ. We compared the time courses over which surface FcγRI became maximal, NADPH oxidase activity was induced, and the antiproliferative effect of IFNγ was detected. Oxidase activity was measured by stimulating cells with PMA or by activating surface FcγRI using aggregated human IgG1 or second antibody crosslinking of mAb 32/Fcγ RI complexes. We found that IFNγ in the absence of additional lymphokines induced high levels of oxidase activity in maximally differentiated U937 cells with even higher levels in the fully differentiated high-Feγ RI expression mutants (>8 nmoles/10 6 cells/min for A12.13 cells). Over the course of differentiation, maximal induced levels of Fcγ RI were reached after 1 to 2 days of IFNγ treatment, prior to the antiproliferative effect of the lymphokine. In contrast, oxidase activity was induced after a lag of approximately 2 days, becoming maximal only after 4 to 6 days of IFNγ treatment. This comparison of the induction of FcγRI with that of oxidase activity triggered through Fcγ RI indicated that the rapid increase of surface receptor was not accompanied by a completion of the pathway of Feγ RI-mediated oxidase activity. However, the time courses of induction detected by PMA and Feγ RI-agonists were coincident suggesting that the development of oxidative capacity could be due to the induction of components required by both the PMA- and surface receptor-mediated pathways. There are several oxidase components that are known to be IFNγ-inducible, such as the oxidase flavoprotein, a b558 cytochrome peptide, and oxidase-requiring cytosolic components, and it is possible that one or a set of these components could be the limiting factor(s) for IFNγ-induced oxidase activity.


Experimental Parasitology | 1981

Toxoplasma gondii: Growth in the absence of host cell protein synthesis

E.R. Pfefferkorn; Lorraine C. Pfefferkorn

Host cell protein synthesis continues when cultured cells are infected by Toxoplasma gondii. In order to determine if this host function is necessary for the parasite we used two independent methods that specifically block cellular protein synthesis. In the first, we infected a temperature-sensitive Chinese hamster ovary cell mutant that has a thermolabile leucyl tRNA synthetase. At the restrictive temperature of 40 C, the mutant cells showed only negligible protein synthesis that was probably mitochondrial. At this temperature, the growth and nucleic acid synthesis of T. gondii proceeded normally and [3H]leucine was specifically incorporated into the parasite as demonstrated by autoradiography. A secpnd method for blocking protein synthesis by the host cell employed treatment of uninfected human fibroblast cells with muconomycin A, an inhibitor of initiation. Repeated washing of monolayer cultures reduced the free muconomycin A to an insignificant level while the cells remained incapable of protein synthesis. T. gondii infected and grew normally in the inhibited cells. Autoradiographic localization of the incorporation of [3H]leucine showed that it was almost exclusively in the intracellular parasites in the cells pretreated with muconomycin A. In the untreated control most of the [3H]leucine was incorporated by the host cell rather than the parasite. We conclude that de novo protein synthesis by the host cell is not required to support the growth of intracellular T. gondii.


Journal of Eukaryotic Microbiology | 1977

Specific Labeling of Intracellular Toxoplasma gondii with Uracil

E. R. Pfefferkorn; Lorraine C. Pfefferkorn


Journal of Biological Chemistry | 1967

Inhibitors of Protein Synthesis in Rat Liver Microsome Fractions

Oscar A. Scornik; Mahlon B. Hoagland; Lorraine C. Pfefferkorn; Elizabeth A. Bishop


Journal of Immunology | 1991

Induction of intracellular Ca2+ mobilization and cytotoxicity by hybrid mouse monoclonal antibodies. Fc gamma RII regulation of Fc gamma RI-triggered functions or signaling?

P. Koolwijk; J. G. J. Van De Winkel; Lorraine C. Pfefferkorn; C.W.M. Jacobs; I. Otten; G. T. Spierenburg; Bert J.E.G. Bast


Journal of Biological Chemistry | 1995

A Novel Role for IgG-Fc TRANSDUCTIONAL POTENTIATION FOR HUMAN HIGH AFFINITY Fcγ RECEPTOR (FcγRI) SIGNALING

Lorraine C. Pfefferkorn; Jan G. J. van de Winkel; Sharon L. Swink


Journal of Biological Chemistry | 1996

INTRACLUSTER RESTRICTION OF FC RECEPTOR GAMMA -CHAIN TYROSINE PHOSPHORYLATION SUBVERTED BY A PROTEIN-TYROSINE PHOSPHATASE INHIBITOR

Lorraine C. Pfefferkorn; Sharon L. Swink

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C.W.M. Jacobs

Radboud University Nijmegen

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