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Tissue cysts is a key survival strategy. The latent bradyzoite form allows the parasite to evade the host immune response while awaiting contact with a new host. Recrudescence of encysted parasites is responsible for most clinical disease and can cause lethal encephalitis in immunocompromised individuals. Therefore, the molecular mechanisms responsible for differentiation between tachyzoites and bradyzoites are of keen interest. Prior work has suggested that bradyzoite differentiation and cell cycle are coupled, with the first detectable initiation of the differentiation program occurring in S/M phase, just prior to mitosis. As bradyzoites mature, theirCroken et al. BMC Genomics 2014, 15:515 http://www.biomedcentral.com/1471-2164/15/Page 6 ofFigure 2 Oocyst development induces genes in common with bradyzoite development. A: Gene sets associated with oocyst sporulation. Fritz et al. assayed the transcriptome of oocysts at days 0, 4, and 10 after being expelled by the feline host [10]. We developed five gene sets based upon day(s) of peak expression as described in the materials and methods. The "early" Acetaminophen gene set was excluded because too many genes (>500) fell into this group. Additional file 1 shows members of the gene sets and the patterns of expression of PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/25751659 oocyst gene sets are shown in Additional file 4. The gradient of color indicates the approximate time of peak expression of the gene sets used (red: early-middle; light green: middle; green: middle-late: blue late; purple: early-late). B: The transcriptome of RH in vitro bradyzoites is enriched for genes associated with middle-stage oocysts. Plotted normalized enrichment scores (NES) for oocyst development gene sets for the datasets reported by Fritz [9]. A positive NES indicates that the gene set is associated with unstressed, tachyzoite parasites, while a negative NES indicates that the gene set is associated with alkaline stressed, in vitro bradyzoites as reported by Lescault [11]. Stars indicate significant enrichment (FWER-adjusted p < 0.05). The color of the bar corresponds to the indicated colors in panel A that depict the approximate time of peak expression of the gene sets used (red: early-middle; light green: middle; green: middle-late: blue late; purple: early-late).metabolism slows. Parasites induced to differentiate in vitro tend to have a slowing of the S/M phase and parasites with dual expression of the bradyzoite marker BAG1 and the tachyzoite marker SAG1 are more likely to be in S/M phase [18]. Induction of bradyzoite markers is observed when tachyzoites are incubated as extracellular tachyzoites for prolonged periods [19], suggesting that these parasites are able to sense alterations in their environment and induce bradyzoite gene expression. Based upon the cell cycle profile of extracellular parasites, this signal is likely to be sensed by parasites in the G1 phase of the cell cycle. Bradyzoite differentiation requires replication [1], but eventually, mature bradyzoites within in vivo tissue cysts complete mitosis and then exit the cell cycle, entering the G0 phase [18]. In addition to greatly slowed metabolism and replication, the other major feature of bradyzoites is cyst formation. During encystation within the host cell, the parasitophorous vacuole is lined with a glycosylated cyst wall, and the parasite expresses a stage-specific antigens, notably BAG1 (bradyzoite antigen 1) and CST1,a glycoprotein present in the bradyzoite cyst wall [20,21]. Detection of these.