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A highly conserved segmental duplication in the subtelomeres of Plasmodium falciparum chromosomes varies in copy number.

Mok BW, Ribacke U, Sherwood E, Wahlgren M - Malar. J. (2008)

Bottom Line: The n-gene transcription levels were found to correlate to the number of n-gene copies.Fragments of SD1 harbouring two or three of the SD1-genes (o-gene, pfmc-2tm, q-gene) were also found in the 3D7 genome.In addition a related second SD, SD2, of approximately 55% sequence identity to SD1 was found duplicated in a fresh clinical isolate but was only present in a single copy in 3D7 and in other P. falciparum lines or clones.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology, Tumor and Cell Biology (MTC), Karolinska Institutet, SE-171 77 Stockholm, Sweden. bobo.mok@ki.se

ABSTRACT

Background: Segmental duplications (SD) have been found in genomes of various organisms, often accumulated at the ends of chromosomes. It has been assumed that the sequence homology in-between the SDs allow for ectopic interactions that may contribute to the emergence of new genes or gene variants through recombinatorial events.

Methods: In silico analysis of the 3D7 Plasmodium falciparum genome, conducted to investigate the subtelomeric compartments, led to the identification of subtelomeric SDs. Sequence variation and copy number polymorphisms of the SDs were studied by DNA sequencing, real-time quantitative PCR (qPCR) and fluorescent in situ hybridization (FISH). The levels of transcription and the developmental expression of copy number variant genes were investigated by qPCR.

Results: A block of six genes of >10 kilobases in size, including var, rif, pfmc-2tm and three hypothetical genes (n-, o- and q-gene), was found duplicated in the subtelomeric regions of chromosomes 1, 2, 3, 6, 7, 10 and 11 (SD1). The number of SD1 per genome was found to vary from 4 to 8 copies in between different parasites. The intragenic regions of SD1 were found to be highly conserved across ten distinct fresh and long-term cultivated P. falciparum. Sequence variation was detected in a approximately 23 amino-acid long hypervariable region of a surface-exposed loop of PFMC-2TM. A hypothetical gene within SD1, the n-gene, encoding a PEXEL/VTS-containing two-transmembrane protein was found expressed in ring stage parasites. The n-gene transcription levels were found to correlate to the number of n-gene copies. Fragments of SD1 harbouring two or three of the SD1-genes (o-gene, pfmc-2tm, q-gene) were also found in the 3D7 genome. In addition a related second SD, SD2, of approximately 55% sequence identity to SD1 was found duplicated in a fresh clinical isolate but was only present in a single copy in 3D7 and in other P. falciparum lines or clones.

Conclusion: Plasmodium falciparum carries multiple sequence conserved SDs in the otherwise highly variable subtelomeres of its chromosomes. The uniqueness of the SDs amongst plasmodium species, and the conserved nature of the genes within, is intriguing and suggests an important role of the SD to P. falciparum.

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Copy numbers of the SD2 in different P. falciparum strains. (A) Ratio based differences (UAM25 over 3D7AH1) of microarray oligonucleotides mapped according to the gene locations on chromosome 1 in 3D7. A black arrow indicates the SD2 found duplicated in UAM25. (B) Genetic organization of the SD2 on the right arm of chromosome 1 in 3D7. Genes found duplicated in UAM25 relative to 3D7, according to CGH data and qPCR, are shown in red. (C) Copy numbers of the SD2 genes in different strains relative to 3D7 parasite confirmed by qPCR.
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Figure 3: Copy numbers of the SD2 in different P. falciparum strains. (A) Ratio based differences (UAM25 over 3D7AH1) of microarray oligonucleotides mapped according to the gene locations on chromosome 1 in 3D7. A black arrow indicates the SD2 found duplicated in UAM25. (B) Genetic organization of the SD2 on the right arm of chromosome 1 in 3D7. Genes found duplicated in UAM25 relative to 3D7, according to CGH data and qPCR, are shown in red. (C) Copy numbers of the SD2 genes in different strains relative to 3D7 parasite confirmed by qPCR.

Mentions: A previous CGH project from this laboratory revealed a subtelomeric gene segment (PFA0685c, PFA0690w and PFA0695c), located on the right end of chromosome 1 in the 3D7 strain, to be duplicated in a fresh clinical isolate (UAM25) [35] (Figure 3A). Further analysis indicates that this locus shares three of the same paralogous genes as SD1s described above, with the same gene order and orientation but with less sequence homology (55% identity). This SD was named SD2. Compared to the eight SD1, SD2 was found to carry the n-gene as a pseudogene and the q-gene (PFA0675w) was found to harbour RESA-like repeats and a DNAJ domain (PFAM database: PF0026; amino acid 1097–1160), which the q-gene of SD1 does not possess. PSI-BLAST analyses of the genes in the SD2 (converged at iteration 3) showed that the q-gene has orthologous genes in P. vivax and in rodent malaria parasites (P. yoelii, P. chabaudi and P. berghei). However, no orthologous genes could be identified for the other SD2 gene-members.


A highly conserved segmental duplication in the subtelomeres of Plasmodium falciparum chromosomes varies in copy number.

Mok BW, Ribacke U, Sherwood E, Wahlgren M - Malar. J. (2008)

Copy numbers of the SD2 in different P. falciparum strains. (A) Ratio based differences (UAM25 over 3D7AH1) of microarray oligonucleotides mapped according to the gene locations on chromosome 1 in 3D7. A black arrow indicates the SD2 found duplicated in UAM25. (B) Genetic organization of the SD2 on the right arm of chromosome 1 in 3D7. Genes found duplicated in UAM25 relative to 3D7, according to CGH data and qPCR, are shown in red. (C) Copy numbers of the SD2 genes in different strains relative to 3D7 parasite confirmed by qPCR.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2279139&req=5

Figure 3: Copy numbers of the SD2 in different P. falciparum strains. (A) Ratio based differences (UAM25 over 3D7AH1) of microarray oligonucleotides mapped according to the gene locations on chromosome 1 in 3D7. A black arrow indicates the SD2 found duplicated in UAM25. (B) Genetic organization of the SD2 on the right arm of chromosome 1 in 3D7. Genes found duplicated in UAM25 relative to 3D7, according to CGH data and qPCR, are shown in red. (C) Copy numbers of the SD2 genes in different strains relative to 3D7 parasite confirmed by qPCR.
Mentions: A previous CGH project from this laboratory revealed a subtelomeric gene segment (PFA0685c, PFA0690w and PFA0695c), located on the right end of chromosome 1 in the 3D7 strain, to be duplicated in a fresh clinical isolate (UAM25) [35] (Figure 3A). Further analysis indicates that this locus shares three of the same paralogous genes as SD1s described above, with the same gene order and orientation but with less sequence homology (55% identity). This SD was named SD2. Compared to the eight SD1, SD2 was found to carry the n-gene as a pseudogene and the q-gene (PFA0675w) was found to harbour RESA-like repeats and a DNAJ domain (PFAM database: PF0026; amino acid 1097–1160), which the q-gene of SD1 does not possess. PSI-BLAST analyses of the genes in the SD2 (converged at iteration 3) showed that the q-gene has orthologous genes in P. vivax and in rodent malaria parasites (P. yoelii, P. chabaudi and P. berghei). However, no orthologous genes could be identified for the other SD2 gene-members.

Bottom Line: The n-gene transcription levels were found to correlate to the number of n-gene copies.Fragments of SD1 harbouring two or three of the SD1-genes (o-gene, pfmc-2tm, q-gene) were also found in the 3D7 genome.In addition a related second SD, SD2, of approximately 55% sequence identity to SD1 was found duplicated in a fresh clinical isolate but was only present in a single copy in 3D7 and in other P. falciparum lines or clones.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology, Tumor and Cell Biology (MTC), Karolinska Institutet, SE-171 77 Stockholm, Sweden. bobo.mok@ki.se

ABSTRACT

Background: Segmental duplications (SD) have been found in genomes of various organisms, often accumulated at the ends of chromosomes. It has been assumed that the sequence homology in-between the SDs allow for ectopic interactions that may contribute to the emergence of new genes or gene variants through recombinatorial events.

Methods: In silico analysis of the 3D7 Plasmodium falciparum genome, conducted to investigate the subtelomeric compartments, led to the identification of subtelomeric SDs. Sequence variation and copy number polymorphisms of the SDs were studied by DNA sequencing, real-time quantitative PCR (qPCR) and fluorescent in situ hybridization (FISH). The levels of transcription and the developmental expression of copy number variant genes were investigated by qPCR.

Results: A block of six genes of >10 kilobases in size, including var, rif, pfmc-2tm and three hypothetical genes (n-, o- and q-gene), was found duplicated in the subtelomeric regions of chromosomes 1, 2, 3, 6, 7, 10 and 11 (SD1). The number of SD1 per genome was found to vary from 4 to 8 copies in between different parasites. The intragenic regions of SD1 were found to be highly conserved across ten distinct fresh and long-term cultivated P. falciparum. Sequence variation was detected in a approximately 23 amino-acid long hypervariable region of a surface-exposed loop of PFMC-2TM. A hypothetical gene within SD1, the n-gene, encoding a PEXEL/VTS-containing two-transmembrane protein was found expressed in ring stage parasites. The n-gene transcription levels were found to correlate to the number of n-gene copies. Fragments of SD1 harbouring two or three of the SD1-genes (o-gene, pfmc-2tm, q-gene) were also found in the 3D7 genome. In addition a related second SD, SD2, of approximately 55% sequence identity to SD1 was found duplicated in a fresh clinical isolate but was only present in a single copy in 3D7 and in other P. falciparum lines or clones.

Conclusion: Plasmodium falciparum carries multiple sequence conserved SDs in the otherwise highly variable subtelomeres of its chromosomes. The uniqueness of the SDs amongst plasmodium species, and the conserved nature of the genes within, is intriguing and suggests an important role of the SD to P. falciparum.

Show MeSH
Related in: MedlinePlus