160 research outputs found
The Enigmatic Role of C9ORF72 in Autophagy
Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterized by the loss of motor neurons resulting in a progressive and irreversible muscular paralysis. Advances in large-scale genetics and genomics have revealed intronic hexanucleotide repeat expansions in the gene encoding C9ORF72 as a main genetic cause of ALS and frontotemporal dementia (FTD), the second most common cause of early-onset dementia after Alzheimer's disease. Novel insights regarding the underlying pathogenic mechanisms of C9ORF72 seem to suggest a synergy of loss and gain of toxic function during disease. C9ORF72, thus far, has been found to be involved in homeostatic cellular pathways, such as actin dynamics, regulation of membrane trafficking, and macroautophagy. All these pathways have been found compromised in the pathogenesis of ALS. In this review, we aim to summarize recent findings on the function of C9ORF72, particularly in the macroautophagy pathway, hinting at a requirement to maintain the fine balance of macroautophagy to prevent neurodegeneration
Antibodies against multiple merozoite surface antigens of the human malaria parasite Plasmodium falciparum inhibit parasite maturation and red blood cell invasion
BACKGROUND: Plasmodium falciparum merozoites expose at their surface a large protein complex, which is composed of fragments of merozoite surface protein 1 (MSP-1; called MSP-183, MSP-130, MSP-138, and MSP-142) plus associated processing products of MSP-6 and MSP-7. During erythrocyte invasion this complex, as well as an integral membrane protein called apical membrane antigen-1 (AMA-1), is shed from the parasite surface following specific proteolysis. Components of the MSP-1/6/7 complex and AMA-1 are presently under development as malaria vaccines. METHODS: The specificities and effects of antibodies directed against MSP-1, MSP-6, MSP-7 on the growth of blood stage parasites were studied using ELISA and the pLDH-assay. To understand the mode of action of these antibodies, their effects on processing of MSP-1 and AMA-1 on the surface of merozoites were investigated. RESULTS: Antibodies targeting epitopes located throughout the MSP-1/6/7 complex interfere with shedding of MSP-1, and as a consequence prevent erythrocyte invasion. Antibodies targeting the MSP-1/6/7 complex have no effect on the processing and shedding of AMA-1 and, similarly, antibodies blocking the shedding of AMA-1 do not affect cleavage of MSP-1, suggesting completely independent functions of these proteins during invasion. Furthermore, some epitopes, although eliciting highly inhibitory antibodies, are only poorly recognized by the immune system when presented in the structural context of the intact antigen. CONCLUSIONS: The findings reported provide further support for the development of vaccines based on MSP-1/6/7 and AMA-1, which would possibly include a combination of these antigens
Antibody responses to the merozoite surface protein-1 complex in cerebral malaria patients in India
<p>Abstract</p> <p>Background</p> <p><it>Plasmodium falciparum </it>infection causes cerebral malaria (CM) in a subset of patients with anti-malarial treatment protecting only about 70% to 80% of patients. Why a subset of malaria patients develops CM complications, including neurological sequelae or death, is still not well understood. It is believed that host immune factors may modulate CM outcomes and there is substantial evidence that cellular immune factors, such as cytokines, play an important role in this process. In this study, the potential relationship between the antibody responses to the merozoite surface protein (MSP)-1 complex (which consists of four fragments namely: MSP-1<sub>83</sub>, MSP-1<sub>30</sub>, MSP-1<sub>38 </sub>and MSP-1<sub>42</sub>), MSP-6<sub>36 </sub>and MSP-7<sub>22 </sub>and CM was investigated.</p> <p>Methods</p> <p>Peripheral blood antibody responses to recombinant antigens of the two major allelic forms of MSP-1 complex, MSP-6<sub>36 </sub>and MSP-7<sub>22 </sub>were compared between healthy subjects, mild malaria patients (MM) and CM patients residing in a malaria endemic region of central India. Total IgG and IgG subclass antibody responses were determined using ELISA method.</p> <p>Results</p> <p>The prevalence and levels of IgG and its subclasses in the plasma varied for each antigen. In general, the prevalence of total IgG, IgG1 and IgG3 was higher in the MM patients and lower in CM patients compared to healthy controls. Significantly lower levels of total IgG antibodies to the MSP-1<sub>f38</sub>, IgG1 levels to MSP-1<sub>d83</sub>, MSP-1<sub>19 </sub>and MSP-6<sub>36 </sub>and IgG3 levels to MSP-1<sub>f42 </sub>and MSP-7<sub>22 </sub>were observed in CM patients as compared to MM patients.</p> <p>Conclusion</p> <p>These results suggest that there may be some dysregulation in the generation of antibody responses to some MSP antigens in CM patients and it is worth investigating further whether perturbations of antibody responses in CM patients contribute to pathogenesis.</p
A multifunctional serine protease primes the malaria parasite for red blood cell invasion
The malaria parasite Plasmodium falciparum replicates within an intraerythrocytic parasitophorous vacuole (PV). Rupture of the host cell allows release (egress) of daughter merozoites, which invade fresh erythrocytes. We previously showed that a subtilisin-like protease called PfSUB1 regulates egress by being discharged into the PV in the final stages of merozoite development to proteolytically modify the SERA family of papain-like proteins. Here, we report that PfSUB1 has a further role in ‘priming' the merozoite prior to invasion. The major protein complex on the merozoite surface comprises three proteins called merozoite surface protein 1 (MSP1), MSP6 and MSP7. We show that just before egress, all undergo proteolytic maturation by PfSUB1. Inhibition of PfSUB1 activity results in the accumulation of unprocessed MSPs on the merozoite surface, and erythrocyte invasion is significantly reduced. We propose that PfSUB1 is a multifunctional processing protease with an essential role in both egress of the malaria merozoite and remodelling of its surface in preparation for erythrocyte invasion
Plasmodium falciparum 19-Kilodalton Merozoite Surface Protein 1 (MSP1)-Specific Antibodies That Interfere with Parasite Growth In Vitro Can Inhibit MSP1 Processing, Merozoite Invasion, and Intracellular Parasite Development
Merozoite surface protein 1 (MSP1) is a target for malaria vaccine development. Antibodies to the 19-kDa carboxy-terminal region referred to as MSP1(19) inhibit erythrocyte invasion and parasite growth, with some MSP1-specific antibodies shown to inhibit the proteolytic processing of MSP1 that occurs at invasion. We investigated a series of antibodies purified from rabbits immunized with MSP1(19) and AMA1 recombinant proteins for their ability to inhibit parasite growth, initially looking at MSP1 processing. Although significant inhibition of processing was mediated by several of the antibody samples, there was no clear relationship with overall growth inhibition by the same antibodies. However, no antibody samples inhibited processing but not invasion, suggesting that inhibition of MSP1 processing contributes to but is not the only mechanism of antibody-mediated inhibition of invasion and growth. Examining other mechanisms by which MSP1-specific antibodies inhibit parasite growth, we show that MSP1(19)-specific antibodies are taken up into invaded erythrocytes, where they persist for significant periods and result in delayed intracellular parasite development. This delay may result from antibody interference with coalescence of MSP1(19)-containing vesicles with the food vacuole. Antibodies raised against a modified recombinant MSP1(19) sequence were more efficient at delaying intracellular growth than those to the wild-type protein. We propose that antibodies specific for MSP1(19) can mediate inhibition of parasite growth by at least three mechanisms: inhibition of MSP1 processing, direct inhibition of invasion, and inhibition of parasite development following invasion. The balance between mechanisms may be modulated by modifying the immunogen used to induce the antibodies
Shift in epitope dominance of IgM and IgG responses to Plasmodium falciparum MSP1 block 4
<p>Abstract</p> <p>Background</p> <p><it>Plasmodium falciparum </it>merozoite surface protein-1 (MSP1) has been extensively studied as a blood-stage malaria vaccine candidate, with most work focused on the conserved 19 kDa and semi-conserved 42 kDa C-terminal regions (blocks 16-17) and the hypervariable N-terminal repeat region (block 2). However, recent genotyping studies suggest that additional regions of MSP1 may be under selective pressure, including a locus of intragenic recombination designated as block 4 within the 3' region of the gene.</p> <p>Methods</p> <p>The current study examined the antibody response to the two parental and two recombinant forms of block 4 and to blocks 16-17 (3D7) in study populations from Colombia, Papua New Guinea and Cameroon that differ in malaria transmission intensity and ethnic composition.</p> <p>Results</p> <p>IgM and IgG antibodies were detected against parental and recombinant MSP1 block 4 peptides in all three populations. Overall, 32-44% of the individuals produced IgM to one or more of the peptides, with most individuals having IgM antibodies reactive with both parental and recombinant forms. In contrast, IgG seropositivity to block 4 varied among populations (range 15-65%), with the majority of antibodies showing specificity for one or a pair of block 4 peptides. The IgG response to block 4 was significantly lower than that to blocks 16-17, indicating block 4 is subdominant. Antibodies to block 4 and blocks 16-17 displayed distinct IgG subclass biases, with block 4 responses biased toward IgG3 and blocks 16-17 toward IgG1. These patterns of responsiveness were consistently observed in the three study populations.</p> <p>Conclusions</p> <p>Production of antibodies specific for each parental and recombinant MSP1 block 4 allele in different populations exposed to <it>P. falciparum </it>is consistent with balancing selection of the MSP1 block 4 region by the immune response of individuals in areas of both low and high malaria transmission. MSP1 block 4 determinants may be important in isolate-specific immunity to <it>P. falciparum</it>.</p
The protein disulfide isomerase ERp57 regulates the steady-state levels of the prion protein
Although the accumulation of a misfolded and protease-resistant form of the prion protein (PrP) is a key event in Prion pathogenesis, the cellular factors involved in its folding and quality control are poorly understood. PrP is a glycosylated and disulfide-bonded protein synthesized at the endoplasmic reticulum (ER). The ER foldase ERp57 (also known as Grp58) is highly expressed in the brain of sporadic and infectious forms of Prion-related disorders. ERp57 is a disulfide isomerase involved in the folding of a subset of glycoproteins in the ER as part of the calnexin/calreticulin cycle. Here we show that levels of ERp57 increase mainly in neurons of Creutzfeldt-Jacob patients. Using gain- and loss-of-function approaches in cell culture we demonstrate that ERp57 expression directly controls the maturation and total levels of wild- type PrP and mutant forms associated with human disease. In addition, we found that PrP physically interacts with ERp57, and also with the closest family member PDIA1, but not ERp72. Furthermore, we generated a conditional knockout mouse for ERp57 in the nervous system and detected a reduction in the steady-state levels of the mono- and non-glycosylated forms of PrP in the brain. In contrast, ERp57 transgenic mice showed increased levels of endogenous PrP. Unexpectedly, ERp57 expression did not affect the susceptibility of cells to ER stress in vitro and in vivo. This study identifies ERp57 as a new modulator of PrP levels and may help understanding the consequences of ERp57 upregulation observed in human disease
Deletion of a Malaria Invasion Gene Reduces Death and Anemia, in Model Hosts
Malaria parasites induce complex cellular and clinical phenotypes, including anemia, cerebral malaria and death in a wide range of mammalian hosts. Host genes and parasite ‘toxins’ have been implicated in malarial disease, but the contribution of parasite genes remains to be fully defined. Here we assess disease in BALB/c mice and Wistar rats infected by the rodent malaria parasite Plasmodium berghei with a gene knock out for merozoite surface protein (MSP) 7. MSP7 is not essential for infection but in P. falciparum, it enhances erythrocyte invasion by 20%. In vivo, as compared to wild type, the P. berghei Δmsp7 mutant is associated with an abrogation of death and a decrease from 3% to 2% in peak, circulating parasitemia. The Δmsp7 mutant is also associated with less anemia and modest increase in the size of follicles in the spleen. Together these data show that deletion of a single parasite invasion ligand modulates blood stage disease, as measured by death and anemia. This work is the first to assess the contribution of a gene present in all plasmodial species in severe disease
Bim Links ER Stress and Apoptosis in Cells Expressing Mutant SOD1 Associated with Amyotrophic Lateral Sclerosis
Endoplasmic reticulum (ER) stress is an important pathway to cell death in amyotrophic lateral sclerosis (ALS). We previously demonstrated that ER stress is linked to neurotoxicity associated with formation of inclusions of mutant Cu,Zn-superoxide dismutase 1 (SOD1). Cells bearing mutant inclusions undergo mitochondrial apoptotic signalling. Here, we demonstrate that the BH3-only protein, Bim, is a direct link between ER stress and mitochondrial apoptosis. In the murine neuroblastoma cell line, Neuro2a, bearing mutant SOD1 inclusions, indicators of both ER stress and apoptosis are expressed. Bim knockdown by siRNA significantly reduced nuclear apoptotic features in these inclusion-bearing cells (but did not affect the proportion of cells overall that bear inclusions). Further, both Bax recruitment to mitochondria and cytochrome c redistribution were also decreased under Bim-depletion conditions. However, upregulation of CHOP, a marker of ER stress, was not reduced by Bim knockdown. Significantly, knockdown of CHOP by siRNA reduced the extent of apoptosis in cells bearing mutant SOD1 inclusions. These sequential links between ER stress, CHOP upregulation, and Bim activation of mitochondrial apoptotic signalling indicate a clear pathway to cell death mediated by mutant SOD1
Altered Prion Protein Expression Pattern in CSF as a Biomarker for Creutzfeldt-Jakob Disease
Creutzfeldt-Jakob disease (CJD) is the most frequent human Prion-related disorder (PrD). The detection of 14-3-3 protein in the cerebrospinal fluid (CSF) is used as a molecular diagnostic criterion for patients clinically compatible with CJD. However, there is a pressing need for the identification of new reliable disease biomarkers. The pathological mechanisms leading to accumulation of 14-3-3 protein in CSF are not fully understood, however neuronal loss followed by cell lysis is assumed to cause the increase in 14-3-3 levels, which also occurs in conditions such as brain ischemia. Here we investigated the relation between the levels of 14-3-3 protein, Lactate dehydrogenase (LDH) activity and expression of the prion protein (PrP) in CSF of sporadic and familial CJD cases. Unexpectedly, we found normal levels of LDH activity in CJD cases with moderate levels of 14-3-3 protein. Increased LDH activity was only observed in a percentage of the CSF samples that also exhibited high 14-3-3 levels. Analysis of the PrP expression pattern in CSF revealed a reduction in PrP levels in all CJD cases, as well as marked changes in its glycosylation pattern. PrP present in CSF of CJD cases was sensitive to proteases. The alterations in PrP expression observed in CJD cases were not detected in other pathologies affecting the nervous system, including cases of dementia and tropical spastic paraparesis/HTLV-1 associated myelopathy (HAM/TSP). Time course analysis in several CJD patients revealed that 14-3-3 levels in CSF are dynamic and show a high degree of variability during the end stage of the disease. Post-mortem analysis of brain tissue also indicated that 14-3-3 protein is upregulated in neuronal cells, suggesting that its expression is modulated during the course of the disease. These results suggest that a combined analysis of 14-3-3 and PrP expression pattern in CSF is a reliable biomarker to confirm the clinical diagnosis of CJD patients and follow disease progression
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