ABSTRACT
The Aedes aegypti mosquito is a vector of many infectious agents, including flaviviruses such as Zika virus. Components of mosquito saliva have pleomorphic effects on the vertebrate host to enhance blood feeding, and these changes also create a favorable niche for pathogen replication and dissemination. Here, we demonstrate that human CD47, which is known to be involved in various immune processes, interacts with a 34-kilodalton mosquito salivary protein named Nest1. Nest1 is up-regulated in blood-fed female A. aegypti and facilitates Zika virus dissemination in human skin explants. Nest1 has a stronger affinity for CD47 than its natural ligand, signal regulatory protein α, competing for binding at the same interface. The interaction between Nest1 with CD47 suppresses phagocytosis by human macrophages and inhibits proinflammatory responses by white blood cells, thereby suppressing antiviral responses in the skin. This interaction elucidates how an arthropod protein alters the human response to promote arbovirus infectivity.
Subject(s)
Aedes , Skin , Zika Virus , Aedes/immunology , Aedes/virology , Animals , Humans , Skin/immunology , Skin/virology , Zika Virus/immunology , Zika Virus/physiology , Female , Insect Proteins/immunology , Zika Virus Infection/immunology , Salivary Proteins and Peptides/immunology , Mosquito Vectors/immunology , Mosquito Vectors/virology , CD47 AntigenABSTRACT
Malaria is initiated as Plasmodium sporozoites are injected into the dermis when an infected mosquito probes on a vertebrate host for a blood meal. Factors in the mosquito saliva, such as AgTRIO, can alter the ability of Anopheles gambiae to transmit Plasmodium. We therefore used CRISPR-Cas9-mediated genome editing to generate AgTRIO knockout (KO) A. gambiae and examined the ability of these mosquitoes to probe on a vertebrate host. AgTRIO KO mosquitoes showed a diminished host probing capacity and required repetitive probing to locate a blood resource to complete a blood meal. This increased probing resulted in enhanced Plasmodium transmission to the vertebrate host. Our data demonstrate the importance of the A. gambiae saliva protein AgTRIO in probing and its influence on the ability of mosquitoes to transmit malaria.
Subject(s)
Anopheles , Animals , Anopheles/parasitology , Anopheles/genetics , Malaria/transmission , Malaria/parasitology , Insect Proteins/genetics , Insect Proteins/metabolism , Mice , CRISPR-Cas Systems/genetics , Female , Mosquito Vectors/parasitology , Mosquito Vectors/geneticsABSTRACT
The Aedes aegypti midgut is crucial for blood digestion, nutrition, reproduction, and pathogen interaction. Using single-cell RNA sequencing, we explored virus infection and transcriptomic changes at the cellular level. We identified 12 distinct cell clusters in the Ae. aegypti midgut post-Zika virus infection, including intestinal stem cells, enteroblasts, enteroendocrine cells (EE), and enterocytes (ECs). The virus was found mainly in specific subsets of ECs and EE. Infection altered transcriptional profiles related to metabolism, signaling, and immune responses. Functional studies highlighted three significantly differentially expressed genes in infected cells. Notably, silencing apolipophorin III reduced virus RNA copy number in the midgut, emphasizing the role of specific genes in viral infection. These findings enhance our understanding of mosquito midgut cell processes during Zika virus infection and suggest potential targets for vector control.
ABSTRACT
Background: Measuring malaria transmission intensity using the traditional entomological inoculation rate is difficult. Antibody responses to mosquito salivary proteins such as SG6 have previously been used as biomarkers of exposure to Anopheles mosquito bites. Here, we investigate four mosquito salivary proteins as potential biomarkers of human exposure to mosquitoes infected with P. falciparum: mosGILT, SAMSP1, AgSAP, and AgTRIO. Methods: We tested population-level human immune responses in longitudinal and cross-sectional plasma samples from individuals with known P. falciparum infection from low and moderate transmission areas in Senegal using a multiplexed magnetic bead-based assay. Results: AgSAP and AgTRIO were the best indicators of recent exposure to infected mosquitoes. Antibody responses to AgSAP, in a moderate endemic area, and to AgTRIO in both low and moderate endemic areas, were significantly higher than responses in a healthy non-endemic control cohort (p-values = 0.0245, 0.0064, and <0.0001 respectively). No antibody responses significantly differed between the low and moderate transmission area, or between equivalent groups during and outside the malaria transmission seasons. For AgSAP and AgTRIO, reactivity peaked 2-4 weeks after clinical P. falciparum infection and declined 3 months after infection. Discussion: Reactivity to both AgSAP and AgTRIO peaked after infection and did not differ seasonally nor between areas of low and moderate transmission, suggesting reactivity is likely reflective of exposure to infectious mosquitos or recent biting rather than general mosquito exposure. Kinetics suggest reactivity is relatively short-lived. AgSAP and AgTRIO are promising candidates to incorporate into multiplexed assays for serosurveillance of population-level changes in P. falciparum-infected mosquito exposure.
ABSTRACT
Malaria begins when an infected mosquito injects saliva containing Plasmodium sporozoites into the skin of a vertebrate host. To prevent malaria, vaccination is the most effective strategy and there is an urgent need for new strategies to enhance current pathogen-based vaccines. Active or passive immunization against a mosquito saliva protein, AgTRIO, contributes to protection against Plasmodium infection of mice. In this study, we generated an AgTRIO mRNA-lipid nanoparticle (LNP) and assessed its potential usefulness as a vaccine against malaria. Immunization of mice with an AgTRIO mRNA-LNP generated a robust humoral response, including AgTRIO IgG2a isotype antibodies that have been associated with protection. AgTRIO mRNA-LNP immunized mice exposed to Plasmodium berghei-infected mosquitoes had markedly reduced initial Plasmodium hepatic infection levels and increased survival compared to control mice. In addition, as the humoral response to AgTRIO waned over 6 months, additional mosquito bites boosted the AgTRIO IgG titers, including IgG1 and IgG2a isotypes, which offers a unique advantage compared to pathogen-based vaccines. These data will aid in the generation of future malaria vaccines that may include both pathogen and vector antigens.
ABSTRACT
Arthropod-borne viruses present important public health challenges worldwide. Viruses such as DENV, ZIKV, and WNV are of current concern due to an increasing incidence and an expanding geographic range, generating explosive outbreaks even in non-endemic areas. The clinical signs associated with infection from these arboviruses are often inapparent, mild, or nonspecific, but occasionally develop into serious complications marked by rapid onset, tremors, paralysis, hemorrhagic fever, neurological alterations, or death. They are predominately transmitted to humans through mosquito bite, during which saliva is inoculated into the skin to facilitate blood feeding. A new approach to prevent arboviral diseases has been proposed by the observation that arthropod saliva facilitates transmission of pathogens. Viruses released within mosquito saliva may more easily initiate host invasion by taking advantage of the host's innate and adaptive immune responses to saliva. This provides a rationale for creating vaccines against mosquito salivary proteins, especially because of the lack of licensed vaccines against most of these viruses. This review aims to provide an overview of the effects on the host immune response by the mosquito salivary proteins and how these phenomena alter the infection outcome for different arboviruses, recent attempts to generate mosquito salivary-based vaccines against flavivirus including DENV, ZIKV, and WNV, and the potential benefits and pitfalls that this strategy involves.
ABSTRACT
CDC-42 regulation of non-muscle myosin/NMY-2 is required for polarity maintenance in the one-cell embryo of Caenorhabditis elegans CDC-42 and NMY-2 regulate polarity throughout embryogenesis, but their contribution to later events of morphogenesis are less understood. We have shown that epidermal enclosure requires the GTPase CED-10/Rac1 and WAVE/Scar complex, its effector, to promote protrusions that drive enclosure through the branch actin regulator Arp2/3. Our analysis here of RGA-8, a homolog of SH3BP1/Rich1/ARHGAP17/Nadrin, with BAR and RhoGAP motifs, suggests it regulates CDC-42, so that actin and myosin/NMY-2 promote ventral enclosure during embryonic morphogenesis. Genetic and molecular data suggest RGA-8 regulates CDC-42, and phenocopies the CDC-42 pathway regulators WASP-1/WSP-1 and the F-BAR proteins TOCA-1 and TOCA-2. Live imaging shows RGA-8 and WSP-1 enrich myosin and regulate F-actin in migrating epidermal cells during ventral enclosure. Loss of RGA-8 alters membrane recruitment of active CDC-42. We propose TOCA proteins and RGA-8 use BAR domains to localize and regenerate CDC-42 activity, thus regulating F-actin levels, through the branched actin regulator WSP-1, and myosin enrichment. RhoGAP RGA-8 thus polarizes epithelia, to promote cell migrations and cell shape changes of embryonic morphogenesis.
Subject(s)
Epithelium/enzymology , Epithelium/metabolism , GTPase-Activating Proteins/metabolism , Morphogenesis/genetics , Organogenesis , Animals , Caenorhabditis elegans/embryology , Caenorhabditis elegans Proteins/metabolism , Cell Cycle Proteins/metabolism , GTP-Binding Proteins/metabolism , Organogenesis/genetics , Signal TransductionABSTRACT
The circadian clock and aging are intertwined. Disruption to the normal diurnal rhythm accelerates aging and corresponds with telomere shortening. Telomere attrition also correlates with increase cellular senescence and incidence of chronic disease. In this report, we examined diurnal association of White Collar 2 (WC-2) in Neurospora and BMAL1 in zebrafish and mice and found that these circadian transcription factors associate with telomere DNA in a rhythmic fashion. We also identified a circadian rhythm in Telomeric Repeat-containing RNA (TERRA), a lncRNA transcribed from the telomere. The diurnal rhythm in TERRA was lost in the liver of Bmal1-/- mice indicating it is a circadian regulated transcript. There was also a BMAL1-dependent rhythm in H3K9me3 at the telomere in zebrafish brain and mouse liver, and this rhythm was lost with increasing age. Taken together, these results provide evidence that BMAL1 plays a direct role in telomere homeostasis by regulating rhythms in TERRA and heterochromatin. Loss of these rhythms may contribute to telomere erosion during aging.
Subject(s)
ARNTL Transcription Factors/physiology , Cellular Senescence , Chromosomes/physiology , Circadian Rhythm , DNA-Binding Proteins/metabolism , Heterochromatin/physiology , Telomere/genetics , Transcription Factors/metabolism , Animals , DNA-Binding Proteins/genetics , Female , Gene Expression Regulation , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Repetitive Sequences, Nucleic Acid , Transcription Factors/geneticsABSTRACT
During embryonic morphogenesis, cells and tissues undergo dramatic movements under the control of F-actin regulators. Our studies of epidermal cell migrations in developing Caenorhabditiselegans embryos have identified multiple plasma membrane signals that regulate the Rac GTPase, thus regulating WAVE and Arp2/3 complexes, to promote branched F-actin formation and polarized enrichment. Here, we describe a pathway that acts in parallel to Rac to transduce membrane signals to control epidermal F-actin through the GTPase RHO-1/RhoA. RHO-1 contributes to epidermal migration through effects on underlying neuroblasts. We identify signals to regulate RHO-1-dependent events in the epidermis. HUM-7, the C. elegans homolog of human MYO9A and MYO9B, regulates F-actin dynamics during epidermal migration. Genetics and biochemistry support that HUM-7 behaves as a GTPase-activating protein (GAP) for the RHO-1/RhoA and CDC-42 GTPases. Loss of HUM-7 enhances RHO-1-dependent epidermal cell behaviors. We identify SAX-3/ROBO as an upstream signal that contributes to attenuated RHO-1 activation through its regulation of HUM-7/Myo9. These studies identify a new role for RHO-1 during epidermal cell migration, and suggest that RHO-1 activity is regulated by SAX-3/ROBO acting on the RhoGAP HUM-7.
Subject(s)
Caenorhabditis elegans Proteins/metabolism , Caenorhabditis elegans/embryology , Caenorhabditis elegans/metabolism , Embryo, Nonmammalian/metabolism , Morphogenesis , rho GTP-Binding Proteins/metabolism , Actins/metabolism , Animals , Axon Guidance , Caenorhabditis elegans/cytology , Cell Cycle Proteins/metabolism , Cell Movement , Embryo, Nonmammalian/cytology , Epidermis/metabolism , Epistasis, Genetic , GTP-Binding Proteins/metabolism , Larva/metabolism , Muscles/metabolism , PhenotypeABSTRACT
The circadian oscillator controls time-of-day gene expression by a network of interconnected feedback loops and is reset by light. The requisite for chromatin regulation in eukaryotic transcription necessitates temporal regulation of histone-modifying and chromatin-remodeling enzymes for proper clock function. CHD1 is known to bind H3K4me3 in mammalian cells, and Neurospora CHD1 is required for proper regulation of the frequency (frq) gene. Based on this, we examined a strain lacking SET1 to determine the role of H3K4 methylation in clock- and light-mediated frq regulation. Expression of frq was altered in strains lacking set1 under both circadian- and light-regulated gene expression. There is a delay in the phasing of H3K4me3 relative to the peak in frq expression. White Collar 2 (WC-2) association with the frq promoter persists longer in Δset1, suggesting a more permissible chromatin state. Surprisingly, SET1 is required for DNA methylation in the frq promoter, indicating a dependence on H3K4me for DNA methylation. The data support a model where SET1 is needed for proper regulation by modulating chromatin at frq.