P falciparum gametocyte proteomes
Ontology highlight
ABSTRACT:
INSTRUMENT(S): LTQ Orbitrap Velos
ORGANISM(S): Plasmodium Falciparum (isolate 3d7)
TISSUE(S): Blood Cell, Blood
DISEASE(S): Plasmodium Falciparum Malaria
SUBMITTER: 
Edwin Lasonder
LAB HEAD: Edwin Lasonder
PROVIDER: PXD004124 | Pride | 2016-07-21
REPOSITORIES: Pride
Publications
Comparative Proteomics and Functional Analysis Reveal a Role of Plasmodium falciparum Osmiophilic Bodies in Malaria Parasite Transmission.
Suárez-Cortés Pablo P Sharma Vikram V Bertuccini Lucia L Costa Giulia G Bannerman Naa-Lamiley NL Sannella Anna Rosa AR Williamson Kim K Klemba Michael M Levashina Elena A EA Lasonder Edwin E Alano Pietro P
Molecular & cellular proteomics : MCP 20160718 10
An essential step in the transmission of the malaria parasite to the Anopheles vector is the transformation of the mature gametocytes into gametes in the mosquito gut, where they egress from the erythrocytes and mate to produce a zygote, which matures into a motile ookinete. Osmiophilic bodies are electron dense secretory organelles of the female gametocytes which discharge their contents during gamete formation, suggestive of a role in gamete egress. Only one protein with no functional annotati ...[more]
Similar Datasets
Project description:For malaria transmission, the parasite must undergo sexual differentiation into mature gametocytes. However, the molecular basis for this critical transition in the parasites life cycle is unknown. Six previously uncharacterized genes, Pfg14.744, Pfg14.745, Pfg14.748, Pfg14.763, Pfg14.752 and Pfg6.6 that are members of a 36 gene Plasmodium falciparum-specific subtelomeric superfamily were found to be expressed in parasites that are committed to sexual development as suggested by co-expression of Pfs16 and Pfg27. Northern blots demonstrated that Pfg14.744 and Pfg14.748 were first expressed before the parasites differentiated into morphologically distinct gametocytes, transcription continued to increase until stage II gametocytes were formed and then rapidly decreased. Immunofluorescence assays indicated that both proteins were only produced in the subpopulation of ring stage parasites that are committed to gametocytogenesis and both localized to the parasitophorous vacuole (PV)b of the early ring stage parasites. As the parasites continued to develop Pfg14.748 remained within the parasitophorous vacuole, while Pfg14.744 was detected in the erythrocyte. The 5' flanking region of either gene alone was sufficient to drive early gametocyte specific expression of green fluorescent protein (GFP). In parasites transfected with a plasmid containing the Pfg14.748 5' flanking region immediately upstream of GFP, fluorescence was observed in a small number of schizonts the cycle before stage I gametocytes were observed. This expression pattern is consistent with commitment to sexual differentiation prior to merozoite release and erythrocyte invasion. Further investigation into the role of these genes in the transition from asexual to sexual differentiation could provide new strategies to block malaria transmission. Microarray analysis was used to compare two clones derived from Plasmodium falciparum strain 3D7 parasites that differ in their ability to undergo gametocytogenesis. Clone G+ produces gametocytes and clone G- produces very few if any gametocytes. RNA was harvested from the cultures when the asexual parasitemia was 0.9-1.48% (day 4) (n=4) after setting up the gametocyte cultures and 5.2-5.58% (day 6) (n=4) prior to the appearance of morphologically distinct gametocytes and used to generate cDNA that was labeled with Cy3 or Cy5 and hybridized to the Plasmodium falciparum 70 mer oligonucleotide microarray developed by DeRisi and co-workers.
2014-10-16 | E-GEOD-62364 | biostudies-arrayexpress
Project description:Sexual differentiation of malaria parasites into gametocytes in the vertebrate host and subsequent gamete fertilisation in mosquitoes is essential for the spreading of the disease. The molecular processes orchestrating these transitions are far from fully understood. Here we report the first transcriptome analysis of male and female Plasmodium falciparum gametocytes coupled with a comprehensive proteome analysis. In male gametocytes there is an enrichment of proteins involved in the formation of flagellated gametes; proteins involved in DNA replication, chromatin organisation and axoneme formation. On the other hand, female gametocytes are enriched in proteins required for zygote formation and functions after fertilisation; protein-, lipid- and energy-metabolism. Integration of transcriptome and proteome data revealed 512 highly expressed maternal transcripts without corresponding protein expression indicating large scale translational repression in P. falciparum female gametocytes for the first time. Despite a high degree of conservation between Plasmodium species, 260 of these ârepressed transcriptsâ have not been previously described. Moreover, for some of these genes, protein expression is only reported in oocysts and sporozoites indicating that repressed transcripts can be partitioned into short- and long-term storage. Finally, these data sets provide an essential resource for identification of vaccine/drug targets and for further mechanistic studies. Examining the transcriptome of p47-mCherry positive female and pDynGFP positive (or double positive) male gametocytes by RNA-seq.
2016-06-28 | E-GEOD-75795 | biostudies-arrayexpress
Project description:Gametocytogenesis and gametogenesis in malaria parasites are complex processes of cell differentiation and development likely involving many gene products. Gametocytes develop in the blood of the vertebrate host but mature gametocytes are not activated until taken up by the mosquito vector. Several distinct mutants have been described that block gametogenesis but the detailed molecular causes for the mutant phenotypes are not understood. To investigate whether a block in gametogenesis also results in a changed transcriptional profile, we studied two gene deletions mutants; act2(-) lacking stage-specific actin II and CDPK4(-) lacking calcium-dependent protein kinase 4. Whole-genome microarray analysis was performed from RNA of mature gametocytes to compare the transcriptomes of the mutants with wild-type Plasmodium berghei. The microarray analysis identified ~12% of all genes being differentially expressed in either or both mutants compared to normal gametocytes, as defined by at least two-fold change in transcript abundance. A large proportion of differentially expressed genes in both mutants overlapped consistent with the developmental gametocyte arrest. Distinct profiles in each mutant were also observed. Microarray experiments were performed as dual-color hybridizations on Agilent-024169 custom whole genome Plasmodium berghei 44K arrays. To compensate for dye-specific effects, a dye-reversal color-swap was applied.
2015-09-15 | E-GEOD-65032 | biostudies-arrayexpress
Project description:Plasmodium falciparum gametocyte stages represent a small fraction of the entire parasite biomass that is present during human malaria infection, yet they alone lead to the transmission of this devastating disease. One of the critical gaps in malaria transmission biology and surveillance is our lack of knowledge about gametocyte biology, especially sexual dimorphic development that may influence transmission from the human to the mosquito. Ratios of male and female gametocytes in the peripheral blood can vary significantly; influenced in part by asexual blood stage and gametocyte density as well as vertebrate and invertebrate host factors. Moreover, the role of sex ratios on gametocyte transmission potential to mosquitoes is unknown and dissecting this process has been hampered by the lack of sex-specific protein markers for the circulating, mature stage V gametocytes. The current evidence suggests a high degree of conservation in gametocyte gene complement across Plasmodium, and therefore presumably for sex-specific genes as well. Therefore, to better our understanding of gametocyte development and subsequent infectiousness to mosquitoes, we undertook a two pronged approach. First, we acquired the mixed, male and female stage V gametocyte proteomes of the NF54 isolate and mature stage V female proteome from Dd2, a strain that is defective in producing mature males. Second, we then undertook a Systematic Subtractive Bioinformatic analysis (filtering) approach to identify sex-specific P. falciparum NF54 protein markers based on a comparison with the Dd2 strain and syntenic male and female proteins from the reanalyzed and updated P. berghei (related rodent malaria parasite) gametocyte proteomes. This has produced a short list of putative 174 male- and 258 female-specific P. falciparum stage V proteins. Furthermore, we generated antibodies against three putative female-specific gametocyte stage V proteins in P. falciparum and confirmed sex-specificity for two proteins and also the loss sex-partitioning for a putative female-specific protein in rodent malaria parasites.
2014-07-25 | PXD000813 | Pride
Project description:During intra-erythrocytic development, late asexually replicating Plasmodium falciparum parasites sequester from peripheral circulation. This facilitates chronic infection and is linked to severe disease and organ-specific pathology including cerebral and placental malaria. Immature gametocytes M-bM-^@M-^S sexual stage precursor cells M-bM-^@M-^S likewise disappear from circulation. Recent work has demonstrated that these sexual stage parasites are located in the hematopoietic system of the bone marrow before mature gametocytes are released into the blood stream to facilitate mosquito transmission. However, as sequestration occurs only in vivo and not during in vitro culture, the mechanisms by which it is regulated and enacted (particularly by the gametocyte stage) remain poorly understood. We generated the most comprehensive P. falciparum functional gene network to date by integrating global transcriptional data from a large set of asexual and sexual in vitro samples, patient-derived in vivo samples, and a new set of in vitro samples profiling sexual commitment. We defined more than 250 functional modules (clusters) of genes that are co-expressed primarily during the intra-erythrocytic parasite cycle, including 35 during sexual commitment and gametocyte development. Comparing the in vivo and in vitro datasets allowed us, for the first time, to map the time point of asexual parasite sequestration in patients to 22 hours post invasion, confirming previous in vitro observations on the dynamics of host cell modification and cytoadherence. Moreover, we were able to define the properties of gametocyte sequestration, demonstrating the presence of two circulating gametocyte populations: gametocyte rings between 0 and ~30 hours post invasion and mature gametocytes after around 7 days post invasion. We used 164/TdTom, a transgenic parasite line expressing a red fluorescent protein reporter under a gametocyte-specific promoter to generate schizont samples. Schizonts were subsequently isolated from both the fluorescent and non-fluorescent population by FACS and prepared for microarray analysis. Two biological replicates were produced for both the fluorescent and the non-fluorescent samples.
2015-01-13 | E-GEOD-64887 | biostudies-arrayexpress
Project description:To identify protein phosphatases (PP) that are essential during asexual blood stage development in Plasmodium berghei, we attempted systematic deletion of the 30 P. berghei PP genes using double homologous recombination. Of these, we performed strand-specific RNA-sequencing (RNA-Seq) analysis for 2 myristoylated PP mutants _ppm2 and _ppm5 lines across relevant lifestages ( activated gametocyte and schizont stages in both _ppm2 and _ppm5 and ookinete stage in _ppm5 alone). Two to four biological replicates each for _ppm2, _ppm5 and wild-type parasites were processed into strand-specific RNA-seq libraries and sequenced in Illumina Hiseq platform using 100-bp paired end chemistry.
2014-04-16 | E-MTAB-2215 | biostudies-arrayexpress
Project description:The identification of Pf377 interacting proteins in the osmiophilic bodies in P. falciparum gametocytes. In a comparative proteomics approach, immunopurified Pfg377 complexes were obtained in wild type and Osmiophilic Body -depleted gametocytes (Pf377 knock out gametocytes) and were analyzed by LC-MS/MS to identify Pfg377 interaction partners in wt gametocytes as candidate for OB associated proteins.
2018-10-26 | PXD004088 | Pride
Project description:RNA-binding proteins, such as DOZI/CITH and Puf2, have been shown to play critical roles in the life cycle in Plasmodium species. One of the characterized functions of these RNA-binding proteins is to bind to mRNAs and regulate their fates, and it is hypothesized that this regulation of mRNA homeostasis of specific transcripts is important for successful infection of both vectors and hosts by these parasites. To further understand the role that other RNA-binding proteins may play in translational repression and other critical processes in the parasite, we investigated ALBA4, another RNA-binding protein implicated in translational repression. We appended a C-terminal GFP-tag to ALBA4 (ALBA4::GFP), and then performed immunoprecipitations on chemically cross-linked samples using Streptavidin-coated beads coated with a biotin-conjugated anti-GFP antibody to capture the ALBA4 complex(es). We performed these experiments in multiple life cycle stages, including sexual stages (gametocytes) and asexual stage (schizonts). In gametocytes, we determined that ALBA4 associates with translational repression machinery. This is also the case in schizonts, however ALBA4 also associates with complexes involved in active translation, mRNA export, and mRNA degradation. We hypothesize that ALBA4 plays a multi-faceted role in mRNA homeostasis by associating with multiple protein complexes in a stage-specific manner.
2017-08-28 | PXD004183 | Pride
Project description:Transcriptional profiling of gametocyte non-producer lines in Plasmodium berghei Transcriptome of gametocyte non producer lines (natural and genetic KO) and parental (820) lines. The aim of the study was to identify key genes involved in the decision to commit to gametocytogenesis in Plasmodium berghei. These microarrays compare naturally selected lines that do not produce gametocytes, and the parental line and additionally a genetic knock out of AP2-G PBANKA_143750. Data published Sinha, Hughes, et, al Nature tbc. 2- colour microarray comparing to common background pool (containing all life cycle stages). Replicates of different life cycle stages of gametocyte non-producer lines and wild tye (WT) parental control lines
2013-12-13 | E-GEOD-53246 | biostudies-arrayexpress
Project description:RNA-binding proteins, such as DOZI/CITH and Puf2, have been shown to play critical roles in the life cycle in Plasmodium species. One of the characterized functions of these RNA-binding proteins is to bind to mRNAs and regulate their fates, and it is hypothesized that this regulation of mRNA homeostasis of specific transcripts is important for successful infection of both vectors and hosts by these parasites. To further understand the role that other RNA-binding proteins may play in translational repression and other critical processes in the parasite, we investigated ALBA4, another RNA-binding protein implicated in translational repression. We appended a C-terminal GFP-tag to ALBA4 (ALBA4::GFP), and then performed immunoprecipitations on chemically cross-linked samples using Streptavidin-coated beads coated with a biotin-conjugated anti-GFP antibody to capture the ALBA4 complex(es). We performed these experiments in multiple life cycle stages, including sexual stages (gametocytes) and asexual stage (schizonts). In gametocytes, we determined that ALBA4 associates with translational repression machinery. This is also the case in schizonts, however ALBA4 also associates with complexes involved in active translation, mRNA export, and mRNA degradation. We hypothesize that ALBA4 plays a multi-faceted role in mRNA homeostasis by associating with multiple protein complexes in a stage-specific manner.
2017-08-28 | PXD006276 | Pride