The obligate intracellular pathogen, Anaplasma phagocytophilum, is the causative agent of life-threatening diseases in humans and animals. A. phagocytophilum is an emerging tick-borne pathogen in the United States, Europe, Africa and Asia, with increasing numbers of infected people and animals every year. It is increasingly recognized that intracellular pathogens modify host cell metabolic pathways to increase infection and transmission in both vertebrate and invertebrate hosts. Recent reports have shown that amino acids are central to the host-pathogen metabolic interaction. In this study, a genome-wide search for components of amino acid metabolic pathways was performed in Ixodes scapularis, the main tick vector of A. phagocytophilum in the United States, for which the genome was recently published. The enzymes involved in the synthesis and degradation pathways of the twenty amino acids were identified. Then, the available transcriptomics and proteomics data was used to characterize the mRNA and protein levels of I. scapularis amino acid metabolic pathway components in response to A. phagocytophilum infection of tick tissues and ISE6 tick cells. Our analysis was focused on the interplay between carbohydrate and amino acid metabolism during A. phagocytophilum infection in ISE6 cells. The results showed that tick cells increase the synthesis of phosphoenolpyruvate (PEP) from tyrosine to control A. phagocytophilum infection. Metabolic pathway analysis suggested that this is achieved by (i) increasing the transcript and protein levels of mitochondrial phosphoenolpyruvate carboxykinase (PEPCK-M), (ii) shunting tyrosine into the tricarboxylic acid (TCA) cycle to increase fumarate and oxaloacetate which will be converted into PEP by PEPCK-M, and (iii) blocking all the pathways that use PEP downstream gluconeogenesis (i.e., de novo serine synthesis pathway (SSP), glyceroneogenesis and gluconeogenesis). While sequestering host PEP may be critical for this bacterium because it cannot actively carry out glycolysis to produce PEP, excess of this metabolite may be toxic for A. phagocytophilum. The present work provides a more comprehensive view of the major amino acid metabolic pathways involved in the response to pathogen infection in ticks, and provides the basis for further studies to develop novel strategies for the control of granulocytic anaplasmosis.

Biologie Moléculaire et Immunologie Parasitaires Ecole Nationale Vétérinaire d'Alfort Université Paris EstMaisons Alfort France

Cell and Molecular Biology Laboratory University of Sao PauloSao Paulo Brazil

Department of Parasitology Faculty of Science University of South BohemiaČeské Budějovice Czechia

Department of Veterinary Pathobiology Center for Veterinary Health Sciences Oklahoma State UniversityStillwater OK United States

Institute of Parasitology Biology Center Czech Academy of SciencesČeské Budějovice Czechia

SaBio Instituto de Investigación en Recursos Cinegéticos IREC Ciudad Real Spain

Zobrazit více v PubMed

Alberdi P., Ayllón N., Cabezas-Cruz A., Bell-Sakyi L., Zweygarth E., Stuen S., et al. (2015). Infection of PubMed DOI

Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. (1990). Basic local alignment search tool. J. Mol. Biol. 215, 403–410. 10.1016/S0022-2836(05)80360-2 PubMed DOI

Amelio I., Cutruzzolá F., Antonov A., Agostini M., Melino G. (2014). Serine and glycine metabolism in cancer. Trends Biochem. Sci. 39, 191–198. 10.1016/j.tibs.2014.02.004 PubMed DOI PMC

Antunes A., Derkaoui M., Terrade A., Denizon M., Deghmane A. E., Deutscher J., et al. (2016). The phosphocarrier protein HPr contributes to meningococcal survival during infection. PLoS ONE 11:e0162434. 10.1371/journal.pone.0162434 PubMed DOI PMC

Ayllón N., Villar M., Galindo R. C., Kocan K. M., Šíma R., López J. A., et al. (2015). Systems biology of tissue-specific response to PubMed DOI PMC

Barabote R. D., Saier M. H., Jr. (2005). Comparative genomic analyses of the bacterial phosphotransferase system. Microbiol. Mol. Biol. Rev. 69, 608–634. 10.1128/MMBR.69.4.608-634.2005 PubMed DOI PMC

Baruch M., Belotserkovsky I., Hertzog B. B., Ravins M., Dov E., McIver K. S., et al. (2014). An extracellular bacterial pathogen modulates host metabolism to regulate its own sensing and proliferation. Cell 156, 97–108. 10.1016/j.cell.2013.12.007 PubMed DOI PMC

Belland R. J., Nelson D. E., Virok D., Crane D. D., Hogan D., Sturdevant D., et al. (2003). Transcriptome analysis of chlamydial growth during IFN-γ-mediated persistence and reactivation. Proc. Natl. Acad. Sci. U.S.A. 100, 15971–15976. 10.1073/pnas.2535394100 PubMed DOI PMC

Berg J. M., Tymoczko J. L., Stryer L. (2002). Biochemistry, 5th Edn New York, NY: W H Freeman Press.

Cabezas-Cruz A., Alberdi P., Ayllón N., Valdés J. J., Pierce R., Villar M., et al. (2016). PubMed DOI PMC

Cabezas-Cruz A., Alberdi P., Valdés J. J., Villar M., de la Fuente J. (2017a). PubMed DOI PMC

Cabezas-Cruz A., Alberdi P., Valdés J. J., Villar M., de la Fuente J. (2017b). Remodeling of tick cytoskeleton in response to infection with PubMed DOI

Cabezas-Cruz A., Estrada-Pe-a A., Rego R. O., de la Fuente J. (2017c). Tick-pathogen ensembles: do molecular interactions lead ecological innovation? Front. Cell. Infect. Microbiol. 7:74. 10.3389/fcimb.2017.00074 PubMed DOI PMC

Chu P., Rodriguez A. R., Arulanandam B. P., Klose K. E. (2011). Tryptophan prototrophy contributes to PubMed DOI PMC

de la Fuente J., Estrada-Peña A., Cabezas-Cruz A., Kocan K. M. (2016a). PubMed DOI

de la Fuente J., Estrada-Peña A., Venzal J. M., Kocan K. M., Sonenshine D. E. (2008). Overview: ticks as vectors of pathogens that cause disease in humans and animals. Front. Biosci. 13:3200. 10.2741/3200 PubMed DOI

de la Fuente J., Torina A., Naranjo V., Caracappa S., Di Marco V., Alongi A., et al. (2005). Infection with PubMed DOI PMC

de la Fuente J., Villar M., Cabezas-Cruz A., Estrada-Pe-a A., Ayllón N., Alberdi P. (2016b). Tick-host-pathogen interactions: conflict and cooperation. PLoS Pathog. 12:e1005488. 10.1371/journal.ppat.1005488 PubMed DOI PMC

de la Fuente J., Waterhouse R. M., Sonenshine D. E., Roe R. M., Ribeiro J. M., Sattelle D. B., et al. (2016c). Tick genome assembled: new opportunities for research on tick-host-pathogen interactions. Front. Cell. Infect. Microbiol. 6:103. 10.3389/fcimb.2016.00103 PubMed DOI PMC

Dunning H. J. C., Lin M., Madupu R., Crabtree J., Angiuoli S. V., Eisen J. A., et al. (2006). Comparative genomics of emerging human ehrlichiosis agents. PLoS Genet. 2:e21. 10.1371/journal.pgen.0020021 PubMed DOI PMC

Finn R. D., Bateman A., Clements J., Coggill P., Eberhardt R. Y., Eddy S. R., et al. (2014). Pfam: the protein families database. Nucleic. Acids. Res. 42, D222-D2230. 10.1093/nar/gkt1223 PubMed DOI PMC

Gaitán S., Tejero C., Ruiz-Amil M. (1983). Some comparative properties of pyruvate kinase in haematopoietic cells and erythrocytes from rainbow trout (Salmo gairdneri R). Comp. Biochem. Physiol. B 74, 801–805. 10.1016/0305-0491(83)90149-9 PubMed DOI

Grüning N. M., Du D., Keller M. A., Luisi B. F., Ralser M. (2014). Inhibition of triosephosphate isomerase by phosphoenolpyruvate in the feedback-regulation of glycolysis. Open Biol. 4:130232. 10.1098/rsob.130232 PubMed DOI PMC

Gulia-Nuss M., Nuss A. B., Meyer J. M., Sonenshine D. E., Roe R. M., Waterhouse R. M., et al. (2016). Genomic insights into the PubMed DOI PMC

Hondalus M. K., Bardarov S., Russell R., Chan J., Jacobs W. R., Jr., Bloom B. R. (2000). Attenuation of and protection induced by a leucine auxotroph of PubMed DOI PMC

Husnik F., Nikoh N., Koga R., Ross L., Duncan R. P., Fujie M., et al. (2013). Horizontal gene transfer from diverse bacteria to an insect genome enables a tripartite nested mealybug symbiosis. Cell 153, 1567–1578. 10.1016/j.cell.2013.05.040 PubMed DOI

Khajanchi B. K., Odeh E., Gao L., Jacobs M. B., Philipp M. T., Lin T., et al. (2015). Phosphoenolpyruvate phosphotransferase system components modulate gene transcription and virulence of PubMed DOI PMC

Kocan K. M., de la Fuente J., Cabezas-Cruz A. (2015). The genus Anaplasma: new challenges after reclassification. Rev. Sci. Tech. 34, 577–586. 10.20506/rst.34.2.2381 PubMed DOI

Kopáček P., Perner J. (2016). Vector Biology: Tyrosine degradation protects blood feeders from death via la grande bouffe. Curr. Biol. 26, R763–R765. 10.1016/j.cub.2016.06.068 PubMed DOI

Leitão-Gonçalves R., Carvalho-Santos Z., Francisco A. P., Fioreze G. T., Anjos M., Baltazar C., et al. (2017). Commensal bacteria and essential amino acids control food choice behavior and reproduction. PLoS Biol. 15:e2000862. 10.1371/journal.pbio.2000862 PubMed DOI PMC

Liu X., Lu R., Xia Y., Sun J. (2010). Global analysis of the eukaryotic pathways and networks regulated by PubMed DOI PMC

Madden T. L., Tatusov R. L., Zhang J. (1996). Applications of network BLAST server. Meth. Enzymol. 266, 131–141. 10.1016/S0076-6879(96)66011-X PubMed DOI

Méndez-Lucas A., Duarte J. A., Sunny N. E., Satapati S., He T., Fu X., et al. (2013). PEPCK-M expression in mouse liver potentiates, not replaces, PEPCK-C mediated gluconeogenesis. J. Hepatol. 59, 105–113. 10.1016/j.jhep.2013.02.020 PubMed DOI PMC

Munderloh U. G., Jauron S. D., Fingerle V., Leitritz L., Hayes S. F., Hautman J. M., et al. (1999). Invasion and intracellular development of the human granulocytic ehrlichiosis agent in tick cell culture. J. Clin. Microbiol. 37, 2518–2524. PubMed PMC

Munderloh U. G., Liu Y., Wang M., Chen C., Kurtti T. J. (1994). Establishment, maintenance and description of cell lines from the tick PubMed DOI

Nye C. K., Hanson R. W., Kalhan S. C. (2008). Glyceroneogenesis is the dominant pathway for triglyceride glycerol synthesis PubMed DOI PMC

O'Callaghan D., Maskell D., Liew F. Y., Easmon C. S., Dougan G. (1988). Characterization of aromatic- and purine-dependent PubMed PMC

Østergaard O., Follmann F., Olsen A. W., Heegaard N. H., Andersen P., Rosenkrands I. (2016). Quantitative protein profiling of PubMed DOI PMC

Olive A. J., Sassetti C. M. (2016). Metabolic crosstalk between host and pathogen: sensing, adapting and competing. Nat. Rev. Microbiol. 14, 221–234. 10.1038/nrmicro.2016.12 PubMed DOI

Owen O. E., Kalhan S. C., Hanson R. W. (2002). The key role of anaplerosis and cataplerosis for citric acid cycle function. J. Biol. Chem. 277, 30409–30412. 10.1074/jbc.R200006200 PubMed DOI

Pfefferkorn E. R. (1984). Interferon gamma blocks the growth of PubMed DOI PMC

Postma P. W., Lengeler J. W., Jacobson G. R. (1993). Phosphoenolpyruvate:carbohydrate phosphotransferase systems of bacteria. Microbiol. Rev. 57, 543–594. PubMed PMC

Samanta D., Semenza G. L. (2016). Serine synthesis helps hypoxic cancer stem cells regulate redox. Cancer Res. 76, 6458–6462. 10.1158/0008-5472.CAN-16-1730 PubMed DOI

Sinclair S. H., Garcia-Garcia J. C., Dumler J. S. (2015). Bioinformatic and mass spectrometry identification of PubMed DOI PMC

Sinclair S. H., Rennoll-Bankert K. E., Dumler J. S. (2014). Effector bottleneck: microbial reprogramming of parasitized host cell transcription by epigenetic remodeling of chromatin structure. Front. Genet. 5:274. 10.3389/fgene.2014.00274 PubMed DOI PMC

Stark R., Pasquel F., Turcu A., Pongratz R. L., Roden M., Cline G. W., et al. (2009). Phosphoenolpyruvate cycling via mitochondrial phosphoenolpyruvate carboxykinase links anaplerosis and mitochondrial GTP with insulin secretion. J. Biol. Chem. 284, 26578–26590. 10.1074/jbc.M109.011775 PubMed DOI PMC

Sterkel M., Perdomo H. D., Guizzo M. G., Barletta A. B., Nunes R. D., Dias F. A., et al. (2016). Tyrosine detoxification is an essential trait in the life history of blood-feeding Arthropods. Curr. Biol. 26, 2188–2193. 10.1016/j.cub.2016.06.025 PubMed DOI

Stuen S., Granquist E. G., Silaghi C. (2013). PubMed DOI PMC

Sunyakumthorn P., Petchampai N., Grasperge B. J., Kearney M. T., Sonenshine D. E., Macaluso K. R. (2013). Gene expression of tissue-specific molecules in PubMed DOI PMC

Villar M., Ayllón N., Alberdi P., Moreno A., Moreno M., Tobes R., et al. (2015). Integrated metabolomics, transcriptomics and proteomics identifies metabolic pathways affected by PubMed DOI PMC

Wang Q., Millet Y. A., Chao M. C., Sasabe J., Davis B. M., Waldor M. K. (2015). A genome-wide screen reveals that the vibrio cholerae phosphoenolpyruvate phosphotransferase system modulates virulence gene expression. Infect. Immun. 83, 3381–3395. 10.1128/IAI.00411-15 PubMed DOI PMC

Wood H., Fehlner-Gardner C., Berry J., Fischer E., Graham B., Hackstadt T., et al. (2003). Regulation of tryptophan synthase gene expression in PubMed DOI

Yang J., Kalhan S. C., Hanson R. W. (2009). What is the metabolic role of phosphoenolpyruvate carboxykinase? J. Biol. Chem. 284, 27025–27029. 10.1074/jbc.R109.040543 PubMed DOI PMC

Yang M., Vousden K. H. (2016). Serine and one-carbon metabolism in cancer. Nat. Rev. Cancer 16, 650–662. 10.1038/nrc.2016.81 PubMed DOI

Zhang Y. J., Rubin E. J. (2013). Feast or famine: the host-pathogen battle over amino acids. Cell. Microbiol. 15, 1079–1087. 10.1111/cmi.12140 PubMed DOI PMC

Zhang Y. J., Reddy M. C., Ioerger T. R., Rothchild A. C., Dartois V., Schuster B. M., et al. (2013). Tryptophan biosynthesis protects mycobacteria from CD4 T-cell-mediated killing. Cell 155, 1296–1308. 10.1016/j.cell.2013.10.045 PubMed DOI PMC

Enlisting the Ixodes scapularis Embryonic ISE6 Cell Line to Investigate the Neuronal Basis of Tick-Pathogen Interactions