A histidine-rich protein gene marks a linkage group favored strongly in a genetic cross of Plasmodium falciparum

Wellems, Thomas E.; Walliker, David; Smith, Cassandra L.; Rosário, Virgílio Estólio do; Maloy, W L; Howard, Russell J.; Carter, Richard; McCutchan, Thomas F.

doi:10.1016/0092-8674(87)90539-3

Cited by 206 publications

(95 citation statements)

References 30 publications

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“…Nucleic Acid Analysis-P. falciparum genomic DNA was purified as described previously (37). Plasmid integration into the 3Ј-UTR of pfcrt was detected by PCR using the primer combinations p6/p7 and p8/p9 (Table I and Fig.…”

Section: Methodsmentioning

confidence: 99%

Chloroquine Resistance Modulated in Vitro by Expression Levels of the Plasmodium falciparum Chloroquine Resistance Transporter

Waller

Muhle

Ursos

et al. 2003

Journal of Biological Chemistry

110

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Plasmodium falciparum malaria is increasingly difficult to treat and control due to the emergence of parasite resistance to the major antimalarials, notably chloroquine. Recent work has shown that the chloroquine resistance phenotype can be conferred by multiple amino acid mutations in the parasite digestive vacuole transmembrane protein PfCRT. Here, we have addressed whether chloroquine resistance can also be affected by changes in expression levels of this protein.Transient transfection reporter assays revealed that truncation of the pfcrt 3-untranslated region just prior to putative polyadenylation sites resulted in a 10-fold decrease in luciferase expression levels. Using allelic exchange on a chloroquine-resistant line (7G8 from Brazil), this truncated 3-untranslated region was inserted downstream of the pfcrt coding sequence, in the place of the endogenous 3-untranslated region. The resulting pfcrt-modified "knockdown" clones displayed a marked decrease in pfcrt transcription and an estimated 30 -40% decrease in PfCRT protein expression levels. [ 3 H]hypoxanthine incorporation assays demonstrated up to a 40% decrease in chloroquine with or without verapamil IC 50 levels of pfcrt knockdown clones, relative to the 7G8 parent. Single-cell photometric analyses were consistent with an altered intracellular pH in the knockdown clones, providing further evidence for a relationship between PfCRT, pH regulation, and chloroquine resistance. Genetic truncation of 3-untranslated regions provides a useful approach for assessing the impact of candidate genes on drug resistance or other quantifiable phenotypes in P. falciparum.

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Section: Methodsmentioning

confidence: 99%

Chloroquine Resistance Modulated in Vitro by Expression Levels of the Plasmodium falciparum Chloroquine Resistance Transporter

Waller

Muhle

Ursos

et al. 2003

Journal of Biological Chemistry

110

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“…During infection of the mosquito host, fertilization between male and female gametes produces a diploid zygote in which meiosis proceeds, allowing recombination between homologous chromosomes and assortment of recombinant chromosomes in haploid progeny. Experimental crosses between laboratory clones of P. falciparum show independent segregation of genes on different chromosomes (2,3). Recombination along each chromosome occurs such that a 1-centiMorgan genetic map distance corresponds to a molecular map distance of approximately 1.5-3.0 ϫ 10 4 bp on average (4).…”

mentioning

confidence: 99%

High recombination rate in natural populations of Plasmodium falciparum

Conway

Roper

Oduola

et al. 1999

Proc. Natl. Acad. Sci. U.S.A.

263

162

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Malaria parasites are sexually reproducing protozoa, although the extent of effective meiotic recombination in natural populations has been debated. If meiotic recombination occurs frequently, compared with point mutation and mitotic rearrangement, linkage disequilibrium between polymorphic sites is expected to decline with increasing distance along a chromosome. The rate of this decline should be proportional to the effective meiotic recombination rate in the population. Multiple polymorphic sites covering a 5-kb region of chromosome 9 (the msp1 gene) have been typed in 547 isolates from six populations in Africa to test for such a decline and estimate its rate in populations of Plasmodium falciparum. The magnitude of two-site linkage disequilibrium declines markedly with increasing molecular map distance between the sites, reaching nonsignificant levels within a map range of 0.3-1.0 kb in five of the populations and over a larger map distance in the population with lowest malaria endemicity. The rate of decline in linkage disequilibrium over molecular map distance is at least as rapid as that observed in most chromosomal regions of other sexually reproducing eukaryotes, such as humans and Drosophila. These results are consistent with the effective recombination rate expected in natural populations of P. falciparum, predicted on the basis of the underlying molecular rate of meiotic crossover and the coefficient of inbreeding caused by self-fertilization events. This is conclusive evidence to reject any hypothesis of clonality or low rate of meiotic recombination in P. falciparum populations. Moreover, the data have major implications for the design and interpretation of population genetic studies of selection on P. falciparum genes.

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“…Subtelomeric deletions in cultured isolates are known to result in the loss of a number of functional genes of Plasmodium falciparum (1)(2)(3)(4)(5)(6)(7). To examine whether this phenomenon occurs in nature or is an artifact of growth in vitro, we used two-dimensional pulsed-field gradient electrophoresis (PFGE) to examine chromosomal DNA from fresh field isolates.…”

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confidence: 99%

Subtelomeric chromosome deletions in field isolates of Plasmodium falciparum and their relationship to loss of cytoadherence in vitro.

Biggs

Kemp

Brown

1989

Proc. Natl. Acad. Sci. U.S.A.

100

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Subtelomeric deletions are responsible for the loss of expression of several Plasmodium falciparum antigens, including the knob-associated histidine-rich protein (KAHRP). Such deletions are detectable by two-dimensional pulsed-field gradient electrophoresis (PFGE) in which the chromosomes separated in dimension 1 are cleaved with Apa I, and the sizes of telomeric fragments are determined in dimension 2. This sensitive technique has enabled us to examine the role of subtelomeric deletions in two aspects of the biology of Plasmodium falciparum. First, we show that similar subtelomeric deletions to those that occur in vitro also occur in field isolates. Second, we demonstrate a correlation between subtelomeric deletions and loss of the phenotype of "cytoadherence" in cultured isolates. Subclones were generated from the cytoadherent cloned isolate ItG2F6, and their phenotypes were examined with respect to cytoadherence, the expression of "knobs," and agglutination of infected erythrocytes with rabbit antiserum. The only chromosomal change detectable by two-dimensional PFGE among subclones that differ from wild type in each of these three characteristics is a deletion of approximately 100 kilobases at one end of chromosome 2. This deletion includes the gene coding for KAHRP and the subtelomeric repeat designated rep20.Subtelomeric deletions in cultured isolates are known to result in the loss of a number of functional genes of Plasmodium falciparum (1-7). To examine whether this phenomenon occurs in nature or is an artifact of growth in vitro, we used two-dimensional pulsed-field gradient electrophoresis (PFGE) to examine chromosomal DNA from fresh field isolates. This technique allows resolution of telomeric fragments after digestion of separated chromosomes with the restriction endonuclease Apa I. It relies on the observation that a conserved complex consisting of telomeric repeats, an Apa I site(s), and a block of rep20 subtelomeric repeats is common to all chromosomes of P. falciparum (Fig.

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A histidine-rich protein gene marks a linkage group favored strongly in a genetic cross of Plasmodium falciparum

Cited by 206 publications

References 30 publications

Chloroquine Resistance Modulated in Vitro by Expression Levels of the Plasmodium falciparum Chloroquine Resistance Transporter

Chloroquine Resistance Modulated in Vitro by Expression Levels of the Plasmodium falciparum Chloroquine Resistance Transporter

High recombination rate in natural populations of Plasmodium falciparum

Subtelomeric chromosome deletions in field isolates of Plasmodium falciparum and their relationship to loss of cytoadherence in vitro.

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