Cassandra Marie Modahl
Email Address
dbscmm@nus.edu.sg
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Publication JNK pathway restricts DENV2, ZIKV and CHIKV infection by activating complement and apoptosis in mosquito salivary glands(Public Library of Science, 2020-08-10) Chowdhury, A.; Modahl, C.M.; Tan, S.T.; Wei Xiang, B.W.; Missé, D.; Vial, T.; Kini, R.M.; Pompon, J.F.; BIOLOGICAL SCIENCES; DUKE-NUS MEDICAL SCHOOLArbovirus infection of Aedes aegypti salivary glands (SGs) determines transmission. However, there is a dearth of knowledge on SG immunity. Here, we characterized SG immune response to dengue, Zika and chikungunya viruses using high-throughput transcriptomics. We also describe a transcriptomic response associated to apoptosis, blood-feeding and lipid metabolism. The three viruses differentially regulate components of Toll, Immune deficiency (IMD) and c-Jun N- terminal Kinase (JNK) pathways. However, silencing of the Toll and IMD pathway components showed variable effects on SG infection by each virus. In contrast, regulation of the JNK pathway produced consistent responses in both SGs and midgut. Infection by the three viruses increased with depletion of the activator Kayak and decreased with depletion of the negative regulator Puckered. Virus-induced JNK pathway regulates the complement factor, Thioester containing protein-20 (TEP20), and the apoptosis activator, Dronc, in SGs. Individual and co-silencing of these genes demonstrate their antiviral effects and that both may function together. Co-silencing either TEP20 or Dronc with Puckered annihilates JNK pathway antiviral effect. Upon infection in SGs, TEP20 induces antimicrobial peptides (AMPs), while Dronc is required for apoptosis independently of TEP20. In conclusion, we revealed the broad antiviral function of JNK pathway in SGs and showed that it is mediated by a TEP20 complement and Dronc-induced apoptosis response. These results expand our understanding of the immune arsenal that blocks arbovirus transmission. © 2020 Chowdhury et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Publication Cysteine-rich secretory proteins (CRISPs) from venomous snakes: An overview of the functional diversity in a large and underappreciated superfamily(MDPI AG, 2020-03-12) Tadokoro, T.; Modahl, C.M.; Maenaka, K.; Aoki-Shioi, N.; BIOLOGICAL SCIENCESThe CAP protein superfamily (Cysteine-rich secretory proteins (CRISPs), Antigen 5 (Ag5), and Pathogenesis-related 1 (PR-1) proteins) is widely distributed, but for toxinologists, snake venom CRISPs are the most familiar members. Although CRISPs are found in the majority of venoms, very few of these proteins have been functionally characterized, but those that have been exhibit diverse activities. Snake venom CRISPs (svCRISPs) inhibit ion channels and the growth of new blood vessels (angiogenesis). They also increase vascular permeability and promote inflammatory responses (leukocyte and neutrophil infiltration). Interestingly, CRISPs in lamprey buccal gland secretions also manifest some of these activities, suggesting an evolutionarily conserved function. As we strive to better understand the functions that CRISPs serve in venoms, it is worth considering the broad range of CRISP physiological activities throughout the animal kingdom. In this review, we summarize those activities, known crystal structures and sequence alignments, and we discuss predicted functional sites. CRISPs may not be lethal or major components of venoms, but given their almost ubiquitous occurrence in venoms and the accelerated evolution of svCRISP genes, these venom proteins are likely to have functions worth investigating. © 2020 by the authors.Publication Proteomic Deep Mining the Venom of the Red-Headed Krait, Bungarus flaviceps(2018) Chapeaurouge, A; Silva, A; Carvalho, P; McCleary, R.J.R; Modahl, C.M; Perales, J; Kini, R.M; Mackessy, S.P; BIOLOGICAL SCIENCESThe use of -omics technologies allows for the characterization of snake venom composition at a fast rate and at high levels of detail. In the present study, we investigated the protein content of Red-headed Krait (Bungarus flaviceps) venom. This analysis revealed a high diversity of snake venom protein families, as evidenced by high-throughput mass spectrometric analysis. We found all six venom protein families previously reported in a transcriptome study of the venom gland of B. flaviceps, including phospholipases A? (PLA?s), Kunitz-type serine proteinase inhibitors (KSPIs), three-finger toxins (3FTxs), cysteine-rich secretory proteins (CRISPs), snaclecs, and natriuretic peptides. A combined approach of automated database searches and de novo sequencing of tandem mass spectra, followed by sequence similarity searches, revealed the presence of 12 additional toxin families. De novo sequencing alone was able to identify 58 additional peptides, and this approach contributed significantly to the comprehensive description of the venom. Abundant protein families comprise 3FTxs (22.3%), KSPIs (19%), acetylcholinesterases (12.6%), PLA?s (11.9%), venom endothelial growth factors (VEGFs, 8.4%), nucleotidases (4.3%), and C-type lectin-like proteins (snaclecs, 3.3%); an additional 11 toxin families are present at significantly lower concentrations, including complement depleting factors, a family not previously detected in Bungarus venoms. The utility of a multifaceted approach toward unraveling the proteome of snake venoms, employed here, allowed detection of even minor venom components. This more in-depth knowledge of the composition of B. flaviceps venom facilitates a better understanding of snake venom molecular evolution, in turn contributing to more effective treatment of krait bites.Publication Venoms of Rear-Fanged Snakes: New Proteins and Novel Activities(Frontiers Media S.A., 2019) Modahl, C.M.; Mackessy, S.P.; BIOLOGICAL SCIENCESSnake venom research has focused on front-fanged venomous snakes because of the high incidence of human morbidity and mortality from envenomations and larger venom yields of these species, while venoms from rear-fanged snakes have been largely neglected. Rear-fanged snakes (RFS) are a phylogenetically diverse collection of species that feed on a variety of prey and show varying prey capture strategies, from constriction to envenomation. In general, RFS venoms share many toxin families with front-fanged snakes, and venoms generally are either a neurotoxic three-finger toxin (3FTx)-dominated venom or an enzymatic metalloproteinase-dominated venom. These venoms have also been discovered to contain several unique venom protein families. New venom protein superfamilies in RFS venoms include matrix metalloproteinases, distinct from but closely related to snake venom metalloproteinases, veficolins, and acid lipases. Specialized three-finger toxins that target select prey taxa have evolved in some RFS venoms, and this prey capture strategy has appeared in multiple RFS species, from Old World Boiga to New World Spilotes and Oxybelis. Though this same protein superfamily is commonly found in the venoms of elapid (front-fanged) snakes, no elapid 3FTxs appear to show prey-specific toxicity (with the exception of perhaps Micrurus). Neofunctionalization of Spilotes sulphureus 3FTx genes has even resulted in the evolution within a single venom of 3FTxs selectively neurotoxic to different prey taxa (mammals or lizards), allowing this non-constricting RFS to take larger mammalian prey. The large number of 3FTx protein sequences available, together with a growing database of RFS venom 3FTxs, make possible predictions concerning structure-function relationships among these toxins and the basis of selective toxicity of specific RFS venom 3FTxs. Rear-fanged snake venoms are therefore of considerable research interest due to the evolutionary novelties they contain, providing insights into the evolution of snake venom proteins and potential predator-prey coevolution in a broader phylogenetic context. Because of the limited complexity of these venoms, they represent a more tractable source to inform about the biological roles of specific venom proteins that are found in the venoms of this rich diversity of snakes. © Copyright © 2019 Modahl and Mackessy.Publication Full-Length Venom Protein cDNA Sequences from Venom-Derived mRNA: Exploring Compositional Variation and Adaptive Multigene Evolution(2016) Modahl C.M.; Mackessy S.P.; BIOLOGICAL SCIENCESEnvenomation of humans by snakes is a complex and continuously evolving medical emergency, and treatment is made that much more difficult by the diverse biochemical composition of many venoms. Venomous snakes and their venoms also provide models for the study of molecular evolutionary processes leading to adaptation and genotype-phenotype relationships. To compare venom complexity and protein sequences, venom gland transcriptomes are assembled, which usually requires the sacrifice of snakes for tissue. However, toxin transcripts are also present in venoms, offering the possibility of obtaining cDNA sequences directly from venom. This study provides evidence that unknown full-length venom protein transcripts can be obtained from the venoms of multiple species from all major venomous snake families. These unknown venom protein cDNAs are obtained by the use of primers designed from conserved signal peptide sequences within each venom protein superfamily. This technique was used to assemble a partial venom gland transcriptome for the Middle American Rattlesnake (Crotalus simus tzabcan) by amplifying sequences for phospholipases A2, serine proteases, C-lectins, and metalloproteinases from within venom. Phospholipase A2 sequences were also recovered from the venoms of several rattlesnakes and an elapid snake (Pseudechis porphyriacus), and three-finger toxin sequences were recovered from multiple rear-fanged snake species, demonstrating that the three major clades of advanced snakes (Elapidae, Viperidae, Colubridae) have stable mRNA present in their venoms. These cDNA sequences from venom were then used to explore potential activities derived from protein sequence similarities and evolutionary histories within these large multigene superfamilies. Venom-derived sequences can also be used to aid in characterizing venoms that lack proteomic profiles and identify sequence characteristics indicating specific envenomation profiles. This approach, requiring only venom, provides access to cDNA sequences in the absence of living specimens, even from commercial venom sources, to evaluate important regional differences in venom composition and to study snake venom protein evolution. ? 2016 Modahl, Mackessy.Publication The RNA binding protein Quaking represses splicing of the Fibronectin EDA exon and downregulates the interferon response(Oxford University Press, 2021-08-24) Liao, Kuo-Chieh; Chuo, Vanessa; Fagg, W. Samuel; Modahl, Cassandra M.; Widen, Steven; Garcia-Blanco, Mariano A.; BIOLOGICAL SCIENCESQuaking (QKI) controls RNA metabolism in many biological processes including innate immunity, where its roles remain incompletely understood. To illuminate these roles, we performed genome scale transcriptome profiling in QKI knockout cells with or without poly(I:C) transfection, a double-stranded RNA analog that mimics viral infection. Analysis of RNA-sequencing data shows that QKI knockout upregulates genes induced by interferons, suggesting that QKI is an immune suppressor. Furthermore, differential splicing analysis shows that QKI primarily controls cassette exons, and among these events, we noted that QKI silences splicing of the extra domain A (EDA) exon in fibronectin (FN1) transcripts. QKI knockout results in elevated production and secretion of FN1-EDA protein, which is a known activator of interferons. Consistent with an upregulation of the interferon response in QKI knockout cells, our results show reduced production of dengue virus-2 and Japanese encephalitis virus in these cells. In conclusion, we demonstrate that QKI downregulates the interferon system and attenuates the antiviral state. © 2021 The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.Publication Transcriptomic signatures of Tacaribe virus-infected Jamaican fruit bats(American Society for Microbiology, 2017) Gerrard, D.L; Hawkinson, A; Sherman, T; Modahl, C.M; Hume, G; Campbell, C.L; Schountz, T; Frietze, S; BIOLOGICAL SCIENCESTacaribe virus (TCRV) is a mammalian arenavirus that was first isolated from artibeus bats in the 1950s. Subsequent experimental infection of Jamaican fruit bats (Artibeus jamaicensis) caused a disease similar to that of naturally infected bats. Although substantial attention has focused on bats as reservoir hosts of viruses that cause human disease, little is known about the interactions between bats and their pathogens. We performed a transcriptome-wide study to illuminate the response of Jamaican fruit bats experimentally infected with TCRV. Differential gene expression analysis of multiple tissues revealed global and organ-specific responses associated with innate antiviral responses, including interferon alpha/beta and Toll-like receptor signaling, activation of complement cascades, and cytokine signaling, among others. Genes encoding proteins involved in adaptive immune responses, such as gamma interferon signaling and costimulation of T cells by the CD28 family, were also altered in response to TCRV infection. Immunoglobulin gene expression was also elevated in the spleens of infected bats, including IgG, IgA, and IgE isotypes. These results indicate an active innate and adaptive immune response to TCRV infection occurred but did not prevent fatal disease. This de novo assembly provides a highthroughput data set of the Jamaican fruit bat and its host response to TCRV infection, which remains a valuable tool to understand the molecular signatures involved in antiviral responses in bats. © 2017 Gerrard et al.Publication High resolution proteomics of Aedes aegypti salivary glands infected with either dengue, Zika or chikungunya viruses identify new virus specific and broad antiviral factors(Nature Research, 2021-12-01) Chowdhury, Avisha; Modahl, Cassandra M.; Missé, Dorothée; Kini, R. Manjunatha; Pompon, Julien; BIOLOGICAL SCIENCES; DUKE-NUS MEDICAL SCHOOLArboviruses such as dengue (DENV), Zika (ZIKV) and chikungunya (CHIKV) viruses infect close to half a billion people per year, and are primarily transmitted through Aedes aegypti bites. Infection-induced changes in mosquito salivary glands (SG) influence transmission by inducing antiviral immunity, which restricts virus replication in the vector, and by altering saliva composition, which influences skin infection. Here, we profiled SG proteome responses to DENV serotype 2 (DENV2), ZIKV and CHIKV infections by using high-resolution isobaric-tagged quantitative proteomics. We identified 218 proteins with putative functions in immunity, blood-feeding or related to the cellular machinery. We observed that 58, 27 and 29 proteins were regulated by DENV2, ZIKV and CHIKV infections, respectively. While the regulation patterns were mostly virus-specific, we separately depleted four uncharacterized proteins that were upregulated by all three viral infections to determine their effects on these viral infections. Our study suggests that gamma-interferon responsive lysosomal thiol-like (GILT-like) has an anti-ZIKV effect, adenosine deaminase (ADA) has an anti-CHIKV effect, salivary gland surface protein 1 (SGS1) has a pro-ZIKV effect and salivary gland broad-spectrum antiviral protein (SGBAP) has an antiviral effect against all three viruses. The comprehensive description of SG responses to three global pathogenic viruses and the identification of new restriction factors improves our understanding of the molecular mechanisms influencing transmission. © 2021, The Author(s).