Characterization and gene expression analysis of the cir multi-gene family of plasmodium chabaudi chabaudi (AS)

Background: The pir genes comprise the largest multi-gene family in Plasmodium, with members found in P. vivax, P. knowlesi and the rodent malaria species. Despite comprising up to 5% of the genome, little is known about the functions of the proteins encoded by pir genes. P. chabaudi causes chronic infection in mice, which may be due to antigenic variation. In this model, pir genes are called cir s and may be involved in this mechanism, allowing evasion of host immune responses. In order to fully understand the role(s) of CIR proteins during P. chabaudi infection, a detailed characterization of the cir gene family was required. Results: The cir repertoire was annotated and a detailed bioinformatic characterization of the encoded CIR proteins was performed. Two major sub-families were identified, which have been named A and B. Members of each sub-family displayed different amino acid motifs, and were thus predicted to have undergone functional divergence. In addition, the expression of the entire cir repertoire was analyzed via RNA sequencing and microarray. Up to 40% of the cir gene repertoire was expressed in the parasite population during infection, and dominant cir transcripts could be identified. In addition, some differences were observed in the pattern of expression between the cir subgroups at the peak of P. chabaudi infection. Finally, specific cir genes were expressed at different time points during asexual blood stages. Conclusions: In conclusion, the large number of cir genes and their expression throughout the intraerythrocytic cycle of development indicates that CIR proteins are likely to be important for parasite survival. In particular, the detection of dominant cir transcripts at the peak of P. chabaudi infection supports the idea that CIR proteins are expressed, and could perform important functions in the biology of this parasite. Further application of the methodologies described here may allow the elucidation of CIR sub-family A and B protein functions, including their contribution to antigenic variation and immune evasion

Concomitant heterochromatinisation and down-regulation of gene expression unveils epigenetic silencing of RELB in an aggressive subset of chronic lymphocytic leukemia in males

<p>Abstract</p> <p>Background</p> <p>The sensitivity of chronic lymphocytic leukemia (CLL) cells to current treatments, both <it>in vitro </it>and <it>in vivo</it>, relies on their ability to activate apoptotic death. CLL cells resistant to DNA damage-induced apoptosis display deregulation of a specific set of genes.</p> <p>Methods</p> <p>Microarray hybridization (Human GeneChip, Affymetrix), immunofluorescent <it>in situ </it>labeling coupled with video-microscopy recording/analyses, chromatin-immunoprecipitation (ChIP), polymerase chain reactions (PCR), real-time quantitative PCR (RT-QPCR) and bisulfite genome sequencing were the main methods applied. Statistical analyses were performed by applying GCRMA and SAM analysis (microarray data) and Student's t-test or Mann & Whitney's U-test.</p> <p>Results</p> <p>Herein we show that, remarkably, in a resistant male CLL cells the vast majority of genes were down-regulated compared with sensitive cells, whereas this was not the case in cells derived from females. This gene down-regulation was found to be associated with an overall gain of heterochromatin as evidenced by immunofluorescent labeling of heterochromatin protein 1α (HP-1), trimethylated histone 3 lysine 9 (3metH3K9), and 5-methylcytidine (5metC). Notably, 17 genes were found to be commonly deregulated in resistant male and female cell samples. Among these, <it>RELB </it>was identified as a discriminatory candidate gene repressed in the male and upregulated in the female resistant cells.</p> <p>Conclusion</p> <p>The molecular defects in the silencing of <it>RELB </it>involve an increase in H3K9- but not CpG-island methylation in the promoter regions. Increase in acetyl-H3 in resistant female but not male CLL samples as well as a decrease of total cellular level of RelB after an inhibition of histone deacetylase (HDAC) by trichostatin A (TSA), further emphasize the role of epigenetic modifications which could discriminate two CLL subsets. Together, these results highlighted the epigenetic <it>RELB </it>silencing as a new marker of the progressive disease in males.</p

Dysfonction télomérique dans les cellules résistantes de leucémie lymphocytaire chronique (LLC)

La LLC est une hémopathie très hétérogène et reste incurable. Les recherches biologiques ont identifié la taille moyenne des télomères (extrémités protectrices des chromosomes) comme un nouveau facteur pronostique (les cellules malignes de la forme grave de la maladie ont des télomères plus courts que celles de la forme indolente).En parallèle, il a été démontré au laboratoire que les cellules malignes de patients LLC peuvent être résistantes (LLC-R) ou sensibles (LLC-S), in vitro, à l apoptose induite par les dommages de l ADN (CDB). Cette différence semble être corrélée à leur sensibilité aux traitements de première ligne et elle est associée à une dérégulation du système de réparation non homologue de l ADN (NHEJ). L objectif de ce travail était de déterminer si la dysfonction télomérique est associée la résistance aux CDBs et de mieux caractériser cette dysfonction dans les cellules de LLC.Les résultats obtenus démontrent que la taille moyenne des télomères est plus courte dans les cellules LLC-R comparés aux LLC-S et que ce raccourcissement est associé à une structure télomérique altérée pouvant limiter l action des systèmes d élongation. Nous avons aussi mis en évidence que la présence de télomères courts dans les cellules LLC-R s accompagne de i) un raccourcissement de la partie télomérique simple brin, ii) la reconnaissance des télomères courts par le système NHEJ et iii) d une perte de signaux télomériques associée à des marqueurs de mauvais pronostic. Ces résultats suggèrent que la dysfonction télomérique est non seulement une caractéristique biologique de la forme grave de la LLC mais pourrait aussi être à l origine de son évolution et/ou de son apparitionPARIS-BIUP (751062107) / SudocSudocFranceF

Epigenetic reprogramming of <i>Plasmodium</i> within the mosquito.

<p>Reset: expression of subtelomeric multigene families is reset in the mosquito by epigenetic reprogramming of the zygote. This ensures that a parasite population will always express all multigene family members from the start of the erythrocytic cycle and gives merozoites the best possible chance of establishing a blood-stage infection every time they emerge from the liver (e.g., it may be beneficial to express all VSA upon liver egress, as a malaria-experienced host will have pre-existing antibodies that recognise a broad repertoire of variant antigens [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1004987#ppat.1004987.ref023" target="_blank">23</a>]). Select and switch: parasites that express (or switch to) multigene family members offering a survival advantage in their new host are retained, whereas parasites that silence these genes are lost through rounds of selection. In time, this leads to a parasite population expressing a narrow repertoire of multigene family members that promotes survival and chronicity. The further into the chronic phase of infection, the better adapted are parasites to their host. <i>Plasmodium</i> virulence therefore increases, and the need to reset gene expression also increases. Reset: in preparation for entry into the next host, all chromatin marks are again erased following gamete fusion. This model of gene expression provides a general mechanism by which all <i>Plasmodium</i> subtelomeric multigene families could be regulated by the mosquito.</p

ICAM-1 is a key receptor mediating cytoadherence and pathology in the Plasmodium chabaudi malaria model

Abstract Background Parasite cytoadherence within the microvasculature of tissues and organs of infected individuals is implicated in the pathogenesis of several malaria syndromes. Multiple host receptors may mediate sequestration. The identity of the host receptor(s), or the parasite ligand(s) responsible for sequestration of Plasmodium species other than Plasmodium falciparum is largely unknown. The rodent malaria parasites may be useful to model interactions of parasite species, which lack the var genes with their respective hosts, as other multigene families are shared between the species. The role of the endothelial receptors ICAM-1 and CD36 in cytoadherence and in the development of pathology was investigated in a Plasmodium chabaudi infection in C57BL/6 mice lacking these receptors. The schizont membrane-associated cytoadherence (SMAC) protein of Plasmodium berghei has been shown to exhibit reduced CD36-associated cytoadherence in P. berghei ANKA-infected mice. Methods Parasite tissue sequestration and the development of acute stage pathology in P. chabaudi infections of mice lacking CD36 or ICAM-1, their respective wild type controls, and in infections with mutant P. chabaudi parasites lacking the smac gene were compared. Peripheral blood parasitaemia, red blood cell numbers and weight change were monitored throughout the courses of infection. Imaging of bioluminescent parasites in isolated tissues (spleen, lungs, liver, kidney and gut) was used to measure tissue parasite load. Results This study shows that neither the lack of CD36 nor the deletion of the smac gene from P. chabaudi significantly impacted on acute-stage pathology or parasite sequestration. By contrast, in the absence of ICAM-1, infected animals experience less anaemia and weight loss, reduced parasite accumulation in both spleen and liver and higher peripheral blood parasitaemia during acute stage malaria. The reduction in parasite tissue sequestration in infections of ICAM-1 null mice is maintained after mosquito transmission. Conclusions These results indicate that ICAM-1-mediated cytoadherence is important in the P. chabaudi model of malaria and suggest that for rodent malarias, as for P. falciparum, there may be multiple host and parasite molecules involved in sequestration

Characterization of the plasmodium interspersed repeats (PIR) proteins of plasmodium chabaudi indicates functional diversity

Plasmodium multigene families play a central role in the pathogenesis of malaria. The Plasmodium interspersed repeat (pir) genes comprise the largest multigene family in many Plasmodium spp. However their function(s) remains unknown. Using the rodent model of malaria, Plasmodium chabaudi, we show that individual CIR proteins have differential localizations within infected red cell (iRBC), suggesting different functional roles in a blood-stage infection. Some CIRs appear to be located on the surface of iRBC and merozoites and are therefore well placed to interact with host molecules. In line with this hypothesis, we show for the first time that a subset of recombinant CIRs bind mouse RBCs suggesting a role for CIR in rosette formation and/or invasion. Together, our results unravel differences in subcellular localization and ability to bind mouse erythrocytes between the members of the cir family, which strongly suggest different functional roles in a blood-stage infection.ASTAR (Agency for Sci., Tech. and Research, S’pore)Published versio