Laboratory of Genomics and Proteomics of Disease Vectors
In our group, we exploit the Ixodes ricinus tick as a model organism to:
- Apply a systems-based and multimodal approach to discover tick effectors determining blood feeding success.
- Understand which vertebrate host proteolytic cascades tick salivary secretions regulate at sites of tick infestation and how they facilitate blood meal uptake and successful completion of the tick lifecycle.
- Test the pharmacological action(s) of recombinant tick salivary proteins and non-coding RNAs (long non coding and microRNAs) and their mechanism of action in the vertebrate host at the molecular, cellular, and organismal level to expedite drug development.
Our research projects aim to:
- Explore basic mechanisms of tick blood feeding success as the conceptual basis for the future development of novel methods/tools (e.g., anti-tick vaccines and diagnostic kits to monitor exposure of vertebrate hosts to arthropod disease vectors) to control tick populations.
- Demonstrate the pharmacological action of tick salivary proteins and non-coding RNAs in the regulation of vertebrate host homeostasis (haemostasis, vascular biology, immunity, neuromodulation) and/or as anti-microbial peptides and test their potential for drug development in the near future.
- Gene overexpression/protein purification
- Biochemistry: enzyme inhibition kinetics-protease inhibitors
- Structural Biology
- Proteomics/Transcriptomics/Non-coding RNAs
- Immune assays: cell-culture assays /animal models
- Tick colony maintenance/tick saliva production
Bensaoud C., Tenzer S., Poplawski A., Medina J.M.,
Mekki I., Voet H., Aparicio-Puerta E.,
Cuveele B., Distler U., Marini F., Hackenberg M.,
Kotsyfakis M. (2022) Quantitative proteomics analysis reveals core and variable tick salivary proteins at the tick-vertebrate host interface Molecular Ecology
Chai J., Chen X., Ye T., Zeng B., Zeng Q., Wu J., Kaščáková B. ,
Martins L., Prudnikova T., Kutá-Smatanová I.,
Kotsyfakis M., Xu X. (2021) Characterization and functional analysis of cathelicidin-MH, a novel frog-derived peptide with anti-septicemic properties eLife
Chávez A.S.O., Wang X., Marnin L., Archer N.K., Hammond H., McClure E.E., Shaw D., Tully B.G., Buskirk A.D., Ford S.L., Butler L.R., Shahi P., Morozova K., Clement C.D., Lawres L., O’ Neal A.J., Ben Mamoun C., Mason K.L., Hobbs B.E., Scoles G., Barry E.M., Sonenshine D.E., Pal U., Valenzuela J.G. , Sztein M.B., Pasetti M.F., Levin M.L.,
Kotsyfakis M., Jay S.M., Huntley J.F., Miller L.S., Pedra J. (2021) Tick extracellular vesicles enable arthropod feeding and promote distinct outcomes of bacterial infection Nature Communications
Bensaoud C., M'ghirbi Y., Faria F., Chmelař J.,
Kotsyfakis M. (2020) Tick salivary compounds for targeted immunomodulatory therapy Frontiers in Immunology
Kotál J., Beránková Z., Kaščáková B. ,
Martins L., Langhansová H., Ederová M., Kutá Smatanová I.,
Kotsyfakis M., Chmelař J. (2021) Iripin-3, a New Salivary Protein Isolated From Ixodes ricinus Ticks, Displays Immunomodulatory and Anti-Hemostatic Properties In Vitro Frontiers in Immunology
Systems biology approaches to better understand disease vector lifecycles
(a) I. ricinus transmits Borrelia burgdorferi (Lyme disease) and several other arboviral diseases via the salivary glands. We used massive de novo sequencing to characterize the transcriptional dynamics of the salivary and midgut tissues of nymphal and adult I. ricinus at various time points after attachment on the vertebrate host (Kotsyfakis et al., 2015, Sci Rep. 5, 9103). A number of gene family members showed stage- and time-specific expression, with 34 candidate histone-modifying proteins suggesting epigenetic control. Midgut transcriptome analysis revealed several enzymes associated with protein, carbohydrate, and lipid digestion, transporters and channels associated with nutrient uptake, and immune-related transcripts including AMPs.
(b) We identified hemocyte transcripts from semi-engorged female ticks by mass sequencing a hemocyte cDNA library and annotating immune-related transcripts based on their hemocyte abundance and their ubiquitous distribution (Kotsyfakis et al., 2015, PLoS Negl Trop Dis. 9, e0003754). 327 transcripts were significantly overexpressed in the hemocyte libraries, including those coding for scavenger receptors, AMPs, pathogen recognition proteins, proteases, protease inhibitors, lipid metabolism, and immune function. The coding sequences deposited to GenBank significantly increased the publicly available dataset now supporting biochemical, pathophysiological, and translational studies. These gene expression analyses (Chmelař et al., 2016a, Trends Parasitol. 32, 242–254) represent a milestone in the field, and the application of quantitative proteomics to ticks with unknown genomes has provided deeper insights into the molecular mechanisms underlying tick-host-pathogen interactions.
Disease vector-host interactions
(a) Given that tick salivary secretions are critical to the success of the tick transmission lifecycle, we systematically reviewed known interactions between tick saliva/salivary gland extracts and the vertebrate immune system (Kotál et al., 2015, J Proteomics. 128, 58–68). This allowed us to promote the hypothesis that tick salivary protein family members display redundancy and pluripotency in their action on host immune responses (Chmelař et al., 2016b, Trends Parasitol. 32, 368–377). This concept is observed in the three major groups of protease inhibitors secreted into saliva: Kunitz inhibitors (anti-hemostatic) and serpins and cystatins (anti-hemostatic and immune system modulators) (Chmelař et al., 2017, Front Cell Infect Microbiol. 7, 216). We described and functionally and structurally characterized the novel immunomodulatory cystatin, Iristatin, in the salivary glands of feeding I. ricinus ticks (Kotál et al., 2019, Cell Mol Life Sci. 76, 2003–2013). Purified recombinant Iristatin inhibited the proteolytic activity of cathepsins, diminished pro-inflammatory cytokine production by different immune cell populations, and inhibited OVA antigen-induced CD4+ T cell proliferation and leukocyte recruitment in vivo. These pluripotent actions may be exploitable as an immunotherapeutic.
(b) We also described novel arthropod miRNAs in I. ricinus salivary glands and saliva (Hackenberg M et al., 2017, RNA. 23, 1259–1269). De novo prediction yielded 67 miRNAs, of which 35 were not present in miRbase, potentially of exosomal origin. These data represent the first direct evidence of tick miRNA-mediated regulation of vertebrate host gene expression at the tick-host interface (Hackenberg M et al., 2017, RNA. 23, 1259–1269), providing candidate miRNAs for drug discovery efforts.
Tripartite interactions between the disease vector/vector-borne pathogens and the host
Noncoding RNAs (ncRNAs) are now known to be (I) transmitted by the vector to possibly modulate vertebrate host responses and (II) regulated in the host by parasites to favor parasite survival. We provided an overview of the involvement of ncRNAs in the parasite-vector-host triad and their effect on host homeostasis (Bensaoud et al., 2019, Trends Parasitol. 35, 715–724). We also reviewed the role that nanovesicles play during pathogen spread, host colonization, and disease pathogenesis (Chávez et al., 2019, J Cell Sci. 132, jcs224212), focusing on a potential role for arthropod exosomes in this tripartite interaction, especially as carriers of long non-coding (lnc)RNAs (Hackenberg and Kotsyfakis, 2018, Trends Parasitol. 34, 549-552).
Towards application development
Two primary interests have driven research into tick salivary secretions: the search for suitable pathogen transmission blocking or "anti-tick" vaccine candidates and the search for novel therapeutics derived from tick salivary components (Chmelař et al., 2019, Front Physiol. 10, 812). We reviewed the major tick salivary protein families exploitable in medical applications (Chmelař et al., 2019, Front Physiol. 10, 812).