Naturally acquired immunity to malaria across the globe varies in intensity and protective powers. Many of the studies on immunity are from hyperendemic regions of Africa. In Asia, particularly in ...India, there are unique opportunities for exploring and understanding malaria immunity relative to host age, co-occurrence of Plasmodium falciparum and Plasmodium vivax infections, varying travel history, and varying disease severity. Variation in immunity in hospital settings is particularly understudied.
A US NIH ICEMR (South Asia) team examined the level of immunity in an Indian malaria patient population visiting or admitted to Goa Medical College and Hospital in Goa, India. Sera from 200 patients of different ages, in different seasons, infected with P. falciparum or P. vivax or both species, and with different clinical severity were applied to an established protein array system with over 1000 P. falciparum and P. vivax antigens. Differential binding of patient IgG to different antigens was measured.
Even though Goa itself has much more P. vivax than P. falciparum, IgG reactivity towards P. falciparum antigens was very strong and comparable to that seen in regions of the world with high P. falciparum endemicity. Of 248 seropositive P. falciparum antigens, the strongest were VAR, MSP10, HSP70, PTP5, AP2, AMA1, and SYN6. In P. vivax patients, ETRAMPs, MSPs, and ApiAP2, sexual stage antigen s16, RON3 were the strongest IgG binders. Both P. falciparum and P. vivax patients also revealed strong binding to new antigens with unknown functions. Seropositives showed antigens unique to the young (HSP40, ACS6, GCVH) or to non-severe malaria (MSP3.8 and PHIST).
Seroreactivity at a major hospital in Southwest India reveals antibody responses to P. falciparum and P. vivax in a low malaria transmission region with much migration. In addition to markers of transmission, the data points to specific leads for possible protective immunity against severe disease. Several, but not all, key antigens overlap with work from different settings around the globe and from other parts of India. Together, these studies confidently help define antigens with the greatest potential chance of universal application for surveillance and possibly for disease protection, in many different parts of India and the world.
The radial distribution of Plasmodium vivax malaria burden has evoked enormous concern among the global research community. In this study, we have investigated the serum proteome alterations in ...non-severe vivax malaria patients before and during patient recuperation starting from the early febrile to the defervescence and convalescent stages of the infection. We have also performed an extensive quantitative proteomics analysis to compare the serum proteome profiles of vivax malaria patients with low (LPVM) and moderately-high (MPVM) parasitemia with healthy community controls. Interestingly, some of the serum proteins such as Serum amyloid A, Apolipoprotein A1, C-reactive protein, Titin and Haptoglobin, were found to be sequentially altered with respect to increased parasite counts. Analysis of a longitudinal cohort of malaria patients indicated reversible alterations in serum levels of some proteins such as Haptoglobin, Apolipoprotein E, Apolipoprotein A1, Carbonic anhydrase 1, and Hemoglobin subunit alpha upon treatment; however, the levels of a few other proteins did not return to the baseline even during the convalescent phase of the infection. Here we present the first comprehensive serum proteomics analysis of vivax malaria patients with different levels of parasitemia and during the acute and convalescent phases of the infection.
During their complex life cycles, the Apicomplexan parasites Plasmodium falciparum and Toxoplasma gondii employ several layers of regulation of their gene expression. One such layer is mediated at ...the level of translation through upstream open reading frames (uORFs). As uORFs are found in the upstream regions of a majority of transcripts in both the parasites, it is essential that their roles in translational regulation be appreciated to a greater extent. This review provides a comprehensive summary of studies that show uORF-mediated gene regulation in these parasites and highlights examples of clinically and physiologically relevant genes, including var2csa in P. falciparum, and ApiAT1 in T. gondii, that exhibit uORF-mediated regulation. In addition to these examples, several studies that use bioinformatics, transcriptomics, proteomics and ribosome profiling also indicate the possibility of widespread translational regulation by uORFs. Further analysis of these genome-wide datasets, taking into account uORFs associated with each gene, will reveal novel genes involved in key biological pathways such as cell-cycle progression, stress-response and pathogenicity. The cumulative evidence from studies presented in this review suggests that uORFs will play crucial roles in regulating gene expression during clinical disease caused by these important human pathogens.
The secretory pathway in
has evolved to transport proteins to the host cell membrane and to an endosymbiotic organelle, the apicoplast. The latter can occur via the ER or the ER-Golgi route. Here, we ...study these three routes using proteins Erythrocyte Membrane Protein-1 (PfEMP1), Acyl Carrier Protein (ACP) and glutathione peroxidase-like thioredoxin peroxidase (PfTPx
) and inhibitors of vesicular transport. As expected, the G protein-dependent vesicular fusion inhibitor AlF
and microtubule destabilizing drug vinblastine block the trafficking of PfEMP-1, a protein secreted to the host cell membrane. However, while both PfTPx
and ACP are targeted to the apicoplast, only ACP trafficking remains unaffected by these treatments. This implies that G protein-dependent vesicles do not play a role in classical apicoplast protein targeting. Unlike the soluble protein ACP, we show that PfTPx
is localized to the outermost membrane of the apicoplast. Thus, the parasite apicoplast acquires proteins via two different pathways: first, the vesicular trafficking pathway appears to handle not only secretory proteins, but an apicoplast membrane protein, PfTPx
; second, trafficking of apicoplast luminal proteins appear to be independent of G protein-coupled vesicles.
Management of severe malaria remains a critical global challenge. In this study, using a multiplexed quantitative proteomics pipeline we systematically investigated the plasma proteome alterations in ...non-severe and severe malaria patients. We identified a few parasite proteins in severe malaria patients, which could be promising from a diagnostic perspective. Further, from host proteome analysis we observed substantial modulations in many crucial physiological pathways, including lipid metabolism, cytokine signaling, complement, and coagulation cascades in severe malaria. We propose that severe manifestations of malaria are possibly underpinned by modulations of the host physiology and defense machinery, which is evidently reflected in the plasma proteome alterations. Importantly, we identified multiple blood markers that can effectively define different complications of severe falciparum malaria, including cerebral syndromes and severe anemia. The ability of our identified blood markers to distinguish different severe complications of malaria may aid in developing new clinical tests for monitoring malaria severity.
Importin α is nuclear transport receptor that recognises nuclear localisation sequences (NLS). The protein has two domains: armadillo (ARM) repeats containing NLS-binding sites and the importin ...β-binding (IBB) domain. The IBB domain mimics an NLS and can bind to the ARM repeats, preventing NLS binding. This phenomenon, called auto-inhibition, is a key regulatory feature for binding and release of NLS-containing cargo by importin α and mutants that lack auto-inhibition show impaired viability in Saccharomyces cerevisiae. The genome of the human malaria parasite, Plasmodium falciparum, contains a single gene for importin α and here we show that the native protein expressed by this gene lacks auto-inhibition, suggesting that P. falciparum parasites possess unconventional mechanisms for regulation of cargo binding and release. Mutation of a single SKR motif (conserved in Plasmodium species) to KRR in P. falciparum importin α restores auto-inhibition. This is the first report of a single-celled eukaryote that has evolved with a single importin α isoform lacking auto-inhibition.
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•P. falciparum contains a single gene for importin α which lacks auto-inhibition.•Mutation of a single motif in P. falciparum importin α restores auto-inhibition.•Auto-inhibition is not crucial for viability of the parasite, unlike in yeast.•The parasite may employ unique strategies for cargo binding and release.•Lack of auto-inhibition may confer advantages to parasites for faster growth.
In Plasmodium vivax malaria, mechanisms that trigger transition from uncomplicated to fatal severe infections are obscure. In this multi-disciplinary study we have performed a comprehensive analysis ...of clinicopathological parameters and serum proteome profiles of vivax malaria patients with different severity levels of infection to investigate pathogenesis of severe malaria and identify surrogate markers of severity. Clinicopathological analysis and proteomics profiling has provided evidences for the modulation of diverse physiological pathways including oxidative stress, cytoskeletal regulation, lipid metabolism and complement cascades in severe malaria. Strikingly, unlike severe falciparum malaria the blood coagulation cascade was not found to be affected adversely in acute P. vivax infection. To the best of our knowledge, this is the first comprehensive proteomics study, which identified some possible cues for severe P. vivax infection. Our results suggest that Superoxide dismutase, Vitronectin, Titin, Apolipoprotein E, Serum amyloid A, and Haptoglobin are potential predictive markers for malaria severity.
The linear syntheses of 4′-C-aminomethyl-2′-O-methyl uridine and cytidine nucleoside phosphoramidites were achieved using glucose as the starting material. The modified RNA building blocks were ...incorporated into small interfering RNAs (siRNAs) by employing solid phase RNA synthesis. Thermal melting studies showed that the modified siRNA duplexes exhibited slightly lower T m (∼1 °C/modification) compared to the unmodified duplex. Molecular dynamics simulations revealed that the 4′-C-aminomethyl-2′-O-methyl modified nucleotides adopt South-type conformation in a siRNA duplex, thereby altering the stacking and hydrogen-bonding interactions. These modified siRNAs were also evaluated for their gene silencing efficiency in HeLa cells using a luciferase-based reporter assay. The results indicate that the modifications are well tolerated in various positions of the passenger strand and at the 3′ end of the guide strand but are less tolerated in the seed region of the guide strand. The modified siRNAs exhibited prolonged stability in human serum compared to unmodified siRNA. This work has implications for the use of 4′-C-aminomethyl-2′-O-methyl modified nucleotides to overcome some of the challenges associated with the therapeutic utilities of siRNAs.
The global burden of malaria and toxoplasmosis has been limited by the use of efficacious anti-parasitic agents, however, emerging resistance in
species and
threatens disease control worldwide, ...implying that new agents/therapeutic targets are urgently needed. Nuclear localization signal (NLS)-dependent transport into the nucleus, mediated by members of the importin (IMP) superfamily of nuclear transporters, has shown potential as a target for intervention to limit viral infection. Here, we show for the first time that IMPα from
and
have promise as targets for small molecule inhibitors. We use high-throughput screening to identify agents able to inhibit
IMPα binding to a
NLS, identifying a number of compounds that inhibit binding in the µM-nM range, through direct binding to
IMPα, as shown in thermostability assays. Of these, BAY 11-7085 is shown to be a specific inhibitor of
IMPα-NLS recognition. Importantly, a number of the inhibitors limited growth by both
and
. The results strengthen the hypothesis that apicomplexan IMPα proteins have potential as therapeutic targets to aid in identifying novel agents for two important, yet neglected, parasitic diseases.
India significantly contributes to the global malaria burden and has the largest population in the world at risk of malaria. This study aims to analyze alterations in the human serum proteome as a ...consequence of non-severe and severe infections by the malaria parasite Plasmodium falciparum to identify markers related to disease severity and to obtain mechanistic insights about disease pathogenesis and host immune responses. In discovery phase of the study, a comprehensive quantitative proteomic analysis was performed using gel-based (2D-DIGE) and gel-free (iTRAQ) techniques on two independent mass spectrometry platforms (ESI-Q-TOF and Q-Exactive mass spectrometry), and selected targets were validated by ELISA. Proteins showing altered serum abundance in falciparum malaria patients revealed the modulation of different physiological pathways including chemokine and cytokine signaling, IL-12 signaling and production in macrophages, complement cascades, blood coagulation, and protein ubiquitination pathways. Some muscle related and cytoskeletal proteins such as titin and galectin-3-binding protein were found to be up-regulated in severe malaria patients. Hemoglobin levels and platelet counts were also found to be drastically lower in severe malaria patients. Identified proteins including serum amyloid A, C-reactive protein, apolipoprotein E and haptoglobin, which exhibited sequential alterations in their serum abundance in different severity levels of malaria, could serve as potential predictive markers for disease severity. To the best of our information, we report here the first comprehensive analysis describing the serum proteomic alterations observed in severe P. falciparum infected patients from different malaria endemic regions of India. This article is part of a Special Issue entitled: Proteomics in India.
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•P. falciparum infection represents the main reason of malaria associated mortality.•First report on proteomic alterations in severe falciparum malaria in Indian population•2D-DIGE and iTRAQ were performed for comparative quantitative proteomic analysis.•SAA, HP, and Apo A1 are potential biomarkers for monitoring malaria severity.•Vital physiological pathways are modulated adversely in severe falciparum malaria.
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