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A HAPPY map of Cryptosporidium parvum.

ABSTRACT: We have constructed a HAPPY map of the apicomplexan parasite Cryptosporidium parvum. We have placed 204 markers on the 10.4-Mb genome, giving an average marker spacing of approximately 50 kb, with an effective resolution of approximately 40 kb. HAPPY mapping (an in vitro linkage technique based on screening approximately haploid amounts of DNA by the polymerase chain reaction) is fast and accurate and is not subject to the distortions inherent in cloning, meiotic recombination, or hybrid cell formation. In addition, little genomic DNA is needed as a substrate, and the AT content of the genome is largely immaterial, making it an ideal method for mapping otherwise intractable parasite genomes. The map, covering all eight chromosomes, consists of 10 linkage groups, each of which has been chromosomally assigned. We have verified the accuracy of the map by several methods, including the construction of a >140-kb PAC contig on chromosome VI. Less than 1% of our markers detect non-rDNA duplicated sequences.

SUBMITTER: Piper MB

PROVIDER: S-EPMC310802 | biostudies-literature | 1998 Dec

REPOSITORIES: biostudies-literature

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A HAPPY map of Cryptosporidium parvum.

Piper M B MB Bankier A T AT Dear P H PH

Genome research 19981201 12

We have constructed a HAPPY map of the apicomplexan parasite Cryptosporidium parvum. We have placed 204 markers on the 10.4-Mb genome, giving an average marker spacing of approximately 50 kb, with an effective resolution of approximately 40 kb. HAPPY mapping (an in vitro linkage technique based on screening approximately haploid amounts of DNA by the polymerase chain reaction) is fast and accurate and is not subject to the distortions inherent in cloning, meiotic recombination, or hybrid cell fo ...[more]

PMID: 9872984

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Project description:BackgroundHundreds of millions of people are infected with cryptosporidiosis annually, with immunocompromised individuals suffering debilitating symptoms and children in socioeconomically challenged regions at risk of repeated infections. There is currently no effective drug available. In order to facilitate the pursuit of anti-cryptosporidiosis targets and compounds, our study spans the classification of the Cryptosporidium parvum kinome and the structural and biochemical characterization of representatives from the CDPK family and a MAP kinase.ResultsThe C. parvum kinome comprises over 70 members, some of which may be promising drug targets. These C. parvum protein kinases include members in the AGC, Atypical, CaMK, CK1, CMGC, and TKL groups; however, almost 35% could only be classified as OPK (other protein kinases). In addition, about 25% of the kinases identified did not have any known orthologues outside of Cryptosporidium spp. Comparison of specific kinases with their Plasmodium falciparum and Toxoplasma gondii orthologues revealed some distinct characteristics within the C. parvum kinome, including potential targets and opportunities for drug design. Structural and biochemical analysis of 4 representatives of the CaMK group and a MAP kinase confirms features that may be exploited in inhibitor design. Indeed, screening CpCDPK1 against a library of kinase inhibitors yielded a set of the pyrazolopyrimidine derivatives (PP1-derivatives) with IC₅₀ values of < 10 nM. The binding of a PP1-derivative is further described by an inhibitor-bound crystal structure of CpCDPK1. In addition, structural analysis of CpCDPK4 identified an unprecedented Zn-finger within the CDPK kinase domain that may have implications for its regulation.ConclusionsIdentification and comparison of the C. parvum protein kinases against other parasitic kinases shows how orthologue- and family-based research can be used to facilitate characterization of promising drug targets and the search for new drugs.

Dataset Information

A HAPPY map of Cryptosporidium parvum.

Publications

A HAPPY map of Cryptosporidium parvum.

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