Project description:Variant antigens that are encoded by large multigene families play an important role in the adaptation and immune evasion of a wide range of pathogens. However, the study of their biological function is significantly hampered by the difficulty in controlling their expression in its cellular setting. The genomes of Plasmodium spp. encode a number of different multigene families that are thought to play a critical role for their survival. However, with the exception of the P. falciparum var genes very little is known about the biological roles of any of the other multigene families. Here we report a highly efficient genetic system to study variant antigens in Plasmodium spp. using the Selection Linked Integration method; we are able to activate the expression of a single member of a multigene of our choice using its endogenous promoter.
Project description:The objective of this study is to provide a novel method to study multigene proteins in Plasmodium spp. The method is based on selection linked integration (SLI), which allows positive selection of genomic integration events. By targeting specific members of multigene families, parasites are being selected for not only genomic integration but also expression of the targeted gene under its endogenous promoter
Project description:The process of erythrocyte invasion by merozoites of Plasmodium falciparum involves multiple steps, including the formation of a moving junction characterized by the redundancy of many of the receptor-ligand interactions involved. Several of the parasite proteins that interact with erythrocyte receptors or participate in other steps of the process of invasion are encoded by small subtelomerically-located multigene families of four to seven members. We report here that members of the multigene families pfRh, eba, rhopH1/clag and acbp exist in either an active or a silenced state. In the case of two members of the rhopH1/clag family, clag3.1 and clag3.2, expression was mutually exclusive. Silencing occurred in the absence of detectable DNA alterations, suggesting that it is transmitted epigenetically. This was unambiguously demonstrated for eba-140, which was silenced by the formation of facultative heterochromatin. Our data demonstrate that variant expression, epigenetic silencing and mutually exclusive expression in Plasmodium are not unique to genes encoding proteins exported to the surface of the erythrocyte like var genes but also occur for genes involved in host cell invasion..
Project description:Plasmodium multigene families are thought to play important roles in the pathogenesis of malaria. Plasmodium interspersed repeat (pir) genes comprise the largest multigene family in many Plasmodium species. However, their expression pattern and localisation remain to be elucidated. Protein subcellular localisation is fundamental to be able to elucidate the functional importance and cell-cell interactions of the PIR proteins. Here, we use the rodent malaria parasite, Plasmodium chabaudi chabaudi, as a model to investigate the localisation pattern of this gene family. We found that most PIR proteins are co-expressed in clusters during acute and chronic infection; members of the S7 clade are predominantly expressed during the acute-phase, whereas members of the L1 clade dominate the chronic-phase of infection. Using peptide antisera specific for S7 or L1 PIRS, we show that S7 and L1 PIRs have different localisations within the infected red blood cells. S7 PIRs are exported into the infected red blood cells cytoplasm where they are co-localised with parasite-induced host cell modifications termed Maurer's clefts, whereas L1 PIRs are localised on or close to the parasitophorous vacuolar membrane. This localisation pattern changes following mosquito transmission and during progression from acute- to chronic-phase of infection. However, neither S7 nor L1 PIR proteins detected by the peptide antisera are localised on the surface of infected red blood cells, suggesting that they are unlikely to be targets of surface variant-specific antibodies or be involved directly in adhesion of infected red blood cells to host cells, as described for Plasmodium falciparum VAR proteins. Their presence on Maurer’s clefts, as seen for Plasmodium falciparum RIFIN and STEVOR proteins, might further suggest trafficking of the PIRs on the surface of the infected erythrocytes. The differences in subcellular localisation of the two major clades of Plasmodium chabaudi PIRs across the blood cycle, and the apparent lack of expression on the red cell surface strongly suggest that the function(s) of this gene family may differ from those of other multigene families of Plasmodium, such as the var genes of Plasmodium falciparum.
Project description:Hypoglycemia is a clinical hallmark of severe malaria, the often-lethal outcome of Plasmodium falciparum infection. Yet, the underlying mechanisms driving the pathogenesis of malaria-associated hypoglycemia remain poorly understood. Here we report that labile heme, an alarmin generated as a byproduct of hemolysis during the blood stage of Plasmodium spp. infection, plays a central role in the development of malaria-associated hypoglycemia. Labile heme recapitulated the hypometabolic response to Plasmodium (chabaudi chabaudi; Pcc) infection in mice, including the development of anorexia, transcriptional repression of hepatic glucose production (HGP) and reduction of glycemia, energy expenditure (EE) as well as core body temperature. While this hypometabolic response is protective against immune-mediated liver damage and anemia, when sustained over time it can lead to hypoglycemia and compromise EE as well as thermoregulation. In response, asexual stages of Plasmodium spp. activate a transcriptional program that reduces virulence in favor of sexual commitment and presumably malaria transmission. In conclusion, malaria-associated hypoglycemia represents a trade-off of a hypometabolic defense strategy against Plasmodium infection.
Project description:Hypoglycemia is a clinical hallmark of severe malaria, the often-lethal outcome of Plasmodium falciparum infection. Yet, the underlying mechanisms driving the pathogenesis of malaria-associated hypoglycemia remain poorly understood. Here we report that labile heme, an alarmin generated as a byproduct of hemolysis during the blood stage of Plasmodium spp. infection, plays a central role in the development of malaria-associated hypoglycemia. Labile heme recapitulated the hypometabolic response to Plasmodium (chabaudi chabaudi; Pcc) infection in mice, including the development of anorexia, transcriptional repression of hepatic glucose production (HGP) and reduction of glycemia, energy expenditure (EE) as well as core body temperature. While this hypometabolic response is protective against immune-mediated liver damage and anemia, when sustained over time it can lead to hypoglycemia and compromise EE as well as thermoregulation. I response, asexual stages of Plasmodium spp. activate a transcriptional program that reduces virulence in favor of sexual commitment and presumably malaria transmission. In conclusion, malaria-associated hypoglycemia represents a trade-off of a hypometabolic defense strategy against Plasmodium infection.
Project description:Transcriptional profiling comparing gut tissue from fifth instar larvae exposed to five different diets, namely cotton fruit or boll (B), cotton flower bud or square (SQ), cotton leaf (L), pinto bean-based artificial diet (PB) and wheat-based artificial Lepidoptera diet (BIO). The generalist lepidopteran herbivore Helicoverpa armigera can consume host plants in more than 40 plant families, and often utilizes several different tissues on the same plant. It is believed that generalists owe their success to the deployment of various members of multigene families of detoxicative and digestive enzymes, a strategy that may also be responsible for rapid evolution of insecticide resistance. However studies of generalist adaptations have been limited to specific genes or gene families, and an overview of how these adaptations are orchestrated at the transcriptional level is lacking. To investigate the previously-shown CBW preferences for different cotton plant structures, we measured net weight gain of larvae that fed on different cotton organs and compared the transcript profiles of larval guts in response to these organs and towards two different artificial diets commonly used for laboratory rearing of this species, using a two-color alternating loop design microarray experiment. Two-color alternating loop design. Biological replicates: 4 (10 individuals per replicate). 20 samples total.
Project description:Transcriptional profiling comparing gut tissue from fifth instar larvae exposed to six different diets, namely cotton fruit or boll (B), tobacco flower bud (TFB), bean pod (BP), chick pea fruit (CKPF), pinto bean-based artificial diet (PB) and wheat-based artificial Lepidoptera diet (BIO). The generalist lepidopteran herbivore Helicoverpa armigera can feed on more than 87 plant species belonging to 48 families. However, life table studies on different crops have revealed that cotton and corn are the most suitable hosts of this pest and tomato, hot pepper and tobacco are suboptimal. It is believed that generalists owe their success to the deployment of various members of multigene families of detoxicative and digestive enzymes, a strategy that may also be responsible for rapid evolution of insecticide resistance. However studies of generalist adaptations have been limited to specific genes or gene families, and an overview of how these adaptations are orchestrated at the transcriptional level is lacking. We compared the transcript profiles of larval guts in response to differentially suitable hosts and towards two different artificial diets commonly used for laboratory rearing of this species, using a two-color alternating loop design microarray experiment. Two-color alternating loop design. Biological replicates: 4 (10 individuals per replicate). 24 samples total.