Cellular immune response to Plasmodium falciparum after pregnancy is related to previous placental infection and parity

BACKGROUND: Malaria in pregnancy is characterised by the sequestration of Plasmodium falciparum-infected erythrocytes in placental intervillous spaces. Placental parasites express a specific phenotype, which allows them to cytoadhere to chondroitin sulfate A expressed by syncytiotrophoblasts. Malaria infection during pregnancy allows the acquisition of antibodies against placental parasites, these antibodies are thought to be involved in protection during subsequent pregnancies. METHODS: To investigate the development of a cellular response to placental parasites during pregnancy, peripheral blood mononuclear cells were collected from women at the time of their confinement. The study was performed in Cameroon where malaria transmission is perennial. In vitro cell proliferation and cytokine production were measured in response to non-malarial activators (concanavalin A and PPD), a recombinant protein from P. falciparum MSP-1, and erythrocytes infected by two P. falciparum lines, RP5 and W2. Like placental parasites, the RP5 line, but not W2, adheres to chondroitin sulfate A and to syncytiotrophoblasts. RESULTS: The proliferative response to all antigens was lower for cells obtained at delivery than 3 months later. Most interestingly, the cellular response to the RP5 line of P. falciparum was closely related to parity. The prevalence rate and the levels of response gradually increased with the number of previous pregnancies. No such relationship was observed with W2 line, or MSP-1 antigen. CONCLUSIONS: This suggests the occurrence of an immune response more specific for the RP5 line in women having had multiple pregnancies, and who are likely to develop immunity to pregnancy-associated parasites. Both humoral and cellular mechanisms may account for the lower susceptibility of multigravidae to malaria

The Fragmented Mitochondrial Ribosomal RNAs of Plasmodium falciparum

The mitochondrial genome in the human malaria parasite Plasmodium falciparum is most unusual. Over half the genome is composed of the genes for three classic mitochondrial proteins: cytochrome oxidase subunits I and III and apocytochrome b. The remainder encodes numerous small RNAs, ranging in size from 23 to 190 nt. Previous analysis revealed that some of these transcripts have significant sequence identity with highly conserved regions of large and small subunit rRNAs, and can form the expected secondary structures. However, these rRNA fragments are not encoded in linear order; instead, they are intermixed with one another and the protein coding genes, and are coded on both strands of the genome. This unorthodox arrangement hindered the identification of transcripts corresponding to other regions of rRNA that are highly conserved and/or are known to participate directly in protein synthesis.The identification of 14 additional small mitochondrial transcripts from P. falciparum and the assignment of 27 small RNAs (12 SSU RNAs totaling 804 nt, 15 LSU RNAs totaling 1233 nt) to specific regions of rRNA are supported by multiple lines of evidence. The regions now represented are highly similar to those of the small but contiguous mitochondrial rRNAs of Caenorhabditis elegans. The P. falciparum rRNA fragments cluster on the interfaces of the two ribosomal subunits in the three-dimensional structure of the ribosome.All of the rRNA fragments are now presumed to have been identified with experimental methods, and nearly all of these have been mapped onto the SSU and LSU rRNAs. Conversely, all regions of the rRNAs that are known to be directly associated with protein synthesis have been identified in the P. falciparum mitochondrial genome and RNA transcripts. The fragmentation of the rRNA in the P. falciparum mitochondrion is the most extreme example of any rRNA fragmentation discovered

SSU rRNA secondary structure.

<p>The <i>P. falciparum</i> mt SSU rRNA secondary structure model is superimposed onto the <i>E. coli</i> SSU rRNA secondary structure model diagram. Regions where <i>P. falciparum</i> has no structure equivalent to <i>E. coli</i> are shown using gray circles and lines. Colored nucleotides compare the <i>P. falciparum</i> sequence to the Three Phylogenetic Domains/Two Organelles (aka 3P2O) consensus sequence from the Gutell lab’s Comparative RNA Web (CRW) Site [URL for CRW Site: <a href="http://www.rna.ccbb.utexas.edu/" target="_blank">http://www.rna.ccbb.utexas.edu/</a>. URL for 16S rRNA conservation diagram: <a href="http://www.rna.ccbb.utexas.edu/SAE/2B/ConsStruc/Diagrams/cons.16.3.3DOM.pdf" target="_blank">http://www.rna.ccbb.utexas.edu/SAE/2B/ConsStruc/Diagrams/cons.16.3.3DOM.pdf</a>]. Upper case colored nucleotides are conserved in at least 98% of the sequences used. For the <i>P. falciparum</i> mt rRNA fragments: red nucleotides match the 3P2O consensus sequence, light blue nucleotides differ, and black nucleotides cannot be compared to the consensus (which is conserved at less than 90%). Base pair symbols are colored when both of the paired nucleotides have the same color. Each fragment is labeled at its 5′ and 3′ ends. Each helix present in <i>P. falciparum</i> is labeled with its helix number in green (<i>e.g.</i>, H500), based on the CRW Site’s helix numbering system [URL: 16S rRNA, <a href="http://www.rna.ccbb.utexas.edu/CAR/1A/Structures/h.16.b.E.coli.hlxnum.pdf" target="_blank">http://www.rna.ccbb.utexas.edu/CAR/1A/Structures/h.16.b.E.coli.hlxnum.pdf</a>]. <b>Inset: </b><i>C. elegans</i> mt SSU rRNA secondary structure model.</p

LSU rRNA secondary structure (3′ half.)

<p>The <i>P. falciparum</i> mt LSU rRNA (3′ half) secondary structure model is superimposed onto the <i>E. coli</i> LSU rRNA (3′ half) secondary structure model diagram. Regions where <i>P. falciparum</i> has no structure equivalent to <i>E. coli</i> are shown using gray circles and lines. Colored nucleotides compare the <i>P. falciparum</i> sequence to the Three Phylogenetic Domains/Two Organelles (aka 3P2O) consensus sequence from the Gutell lab’s Comparative RNA Web (CRW) Site [URL for CRW Site: <a href="http://www.rna.ccbb.utexas.edu/" target="_blank">http://www.rna.ccbb.utexas.edu/</a>. URL for 23S rRNA (3′ half) conservation diagram: <a href="http://www.rna.ccbb.utexas.edu/SAE/2B/ConsStruc/Diagrams/cons.23.3.3DOM.3.pdf" target="_blank">http://www.rna.ccbb.utexas.edu/SAE/2B/ConsStruc/Diagrams/cons.23.3.3DOM.3.pdf</a>]. Upper case colored nucleotides are conserved in at least 98% of the sequences used. For the <i>P. falciparum</i> mt rRNA fragments: red nucleotides match the 3P2O consensus sequence, light blue nucleotides differ, and black nucleotides cannot be compared to the consensus (which is conserved at less than 90%). Base pair symbols are colored when both of the paired nucleotides have the same color. Each fragment is labeled at its 5′ and 3′ ends. Each helix present in <i>P. falciparum</i> is labeled with its helix number in green (<i>e.g.</i>, H500), based on the CRW Site’s helix numbering system [URL: 23S rRNA 3′ half, <a href="http://www.rna.ccbb.utexas.edu/CAR/1A/Structures/h.233.b.E.coli.hlxnum.pdf" target="_blank">http://www.rna.ccbb.utexas.edu/CAR/1A/Structures/h.233.b.E.coli.hlxnum.pdf</a>]. <b>Inset</b>: <i>C. elegans</i> mt LSU rRNA (3′ half) secondary structure model.</p

rRNA tertiary structure.

<p>The <i>P. falciparum</i> mt rRNAs are superimposed on a space-filling model of the three dimensional rRNA structure. Each individual <i>P. falciparum</i> rRNA fragment is colored and labeled; regions of the model with no <i>P. falciparum</i> equivalent are colored gray. Functional regions of the rRNAs are labeled in black. Full-page versions of each panel are available as <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038320#pone.0038320.s012" target="_blank">Figures S12</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038320#pone.0038320.s016" target="_blank">S16</a>. (<b>A</b>) <i>P. falciparum</i> SSU rRNA superimposed on <i>Thermus thermophilus</i> (PDB ID 1J5E; left  =  front/interface side, right  =  back); (<b>B</b>) <i>P. falciparum</i> LSU rRNA superimposed on <i>Haloarcula marismortui</i> (PDB ID 1S72; left  =  crown/interface side, right  =  back); (<b>C–E</b>) secondary structure diagrams for SSU, 5′ LSU, and 3′ LSU rRNAs, respectively, with each <i>P. falciparum</i> mt rRNA fragment color-coordinated with the fragment colors in the three-dimensional structure.</p

Schematic map of the <i>P. falciparum</i> mt genome.

<p>A schematic map of the 6 kb element is shown, with genes above or below the line depending on direction of transcription (orange arrows: left to right above the line and right to left below). Because the <i>P. falciparum</i> mt genome is tandemly repeated, the endpoints shown are the endpoints of Genbank submission M76611, rather than actual structure. Protein coding genes are indicated by white boxes, small mt rDNA sequences are shown with blue (SSU rRNA) and green (LSU rRNA) boxes and labels, and the locations of RNA23t-RNA27t are indicated with red arrows and labels. Gene abbreviations: <i>cox</i>1 and <i>cox</i>3, cytochrome <i>c</i> oxidase subunits I and III; <i>cob</i>, cytochrome <i>b</i>; LA-LG, LSU rRNA fragments; SA-SF, SSU rRNA fragments; 1–22, RNA1-RNA22; 23t-27t, RNA23t-RNA27t.</p

<i>P. falciparum</i> small mt RNA mapping and identification.

a<p>Naming convention reflects history of identification <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038320#pone.0038320-Feagin3" target="_blank">[4]</a>. RNA23t through RNA27t are tentative assignments. We have not detected them in RNA blotting experiments, but sequence conservation and comparative abundance of cDNAs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038320#pone.0038320-Raabe1" target="_blank">[29]</a> suggest they are mature RNAs or abundant processing products.</p>b<p>5′ ends are based on primer extension data, except for RNA1 (RNase protection), RNAs 9 and 14–22 (5′ RACE), and RNA21 (Raabe, <i>et al.</i><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038320#pone.0038320-Raabe1" target="_blank">[29]</a> cDNAs and sequence conservation). 3′ ends were determined by 3′ RACE except for SSUE, LSUB, and LSUC (inferred from 5′ end location and transcript size). We have confirming RNase protection data for many of the 5′ and 3′ ends. *, coordinates inferred.</p>c<p>cDNAs corresponding to mapped <i>P. falciparum</i> mt RNAs. Each designation is name/number of cDNAs.</p>d<p>The observed size from denaturing polyacrylamide gels is noted for the RNA size, compared to the gene size (DNA) based on the location of 5′ and 3′ ends. ?, RNA identified by RACE, not blotting.</p>e<p>Assignment of <i>P. falciparum</i> small mt RNAs to large and small subunit rRNAs; numbers indicate linear order relative to conventional rRNAs.</p