Our lab is interested in the molecular events that enable apicomplexan parasites to remain widespread and deadly infectious agents. These single-celled eukaryotes comprise a phylum of organisms that parasitize diverse animal hosts. Many important human pathogens belong to this group, including the causative agents of malaria (Plasmodium spp.), cryptosporidiosis (Cryptosporidium spp.), and toxoplasmosis (Toxoplasma gondii). We use T. gondii to model features conserved throughout the phylum, such as their reliance on calcium signaling to regulate motility. We combine several approaches that span phospho-proteomics, chemical-genetics, and genome editing to investigate the unique biology of these organisms. Our work seeks to expand our understanding of eukaryotic diversity and identify specific features that can be targeted to treat parasite infections. For more information on our work, visit iBiology.org for Sebastian’s lectures on Toxoplasma and other stories of cutting-edge research in the life sciences, available for free to a global audience.
HiT the Toxoplasma genome
April 28, 2022. Tyler’s paper on the high-throughput tagging (HiT) of Toxoplasma genes is now published in Nature Microbiology. The paper describes a new method of CRISPR-based screening that manipulates genes through tagging instead of disrupting. Using this approach, we discovered a new regulator of the parasite lytic cycle.
Dr. Tyler A. Smith
April 14, 2022. Tyler successfully defends his thesis on the High-throughput functionalization of the Toxoplasma kinome. Congratulations Dr. Smith!
Dylan receives NSF fellowship
April 14, 2022. Congratulations to Dylan McCormick for receiving one of the National Science Foundation Graduate Research Fellowship Program awards!
Changes in cytosolic calcium regulate eukaryotic cellular responses as diverse as membrane repair and muscle contraction. In apicomplexan parasites, calcium regulates motility in part through the regulation of adhesin exocytosis and myosin-motor function. We seek to understand the molecular details of these processes, examining both the events that lead to cytosolic calcium changes and the signaling events that follow them.
We are interested in how calcium signals are decoded by protein kinases, and in particular the role of calcium-dependent protein kinases (CDPKs) as the primary calcium-responsive kinases in parasites. Our work, and that of many others, has defined diverse roles for these kinases in regulating important events during the life cycle of apicomplexans. We have recently used alpaca-derived single-domain antibodies to probe the structure of CDPKs, defining a new mode of allosteric inhibition. We continue to develop biochemical methods to study these kinases, in addition to chemical-genetic approaches to study their function in vivo.
We have engineered a panel of strains to study individual CDPKs. This approach relies on mutating the “gatekeeper” residue that restricts the depth of the ATP-binding pocket. This allows us to specifically inhibit or identify the targets of individual kinases. Using this approach, we have previously elucidated the distinct roles of TgCDPK1 and TgCDPK3 during the T. gondii lytic cycle. We continue to investigate the functions of these and other CDPKs, relying in part on phospho-proteomic methods to identify specific kinase targets.
Much of our work is made possible through a variety of genome-engineering methods. We are interested in developing new methods to enable efficient functional analysis of parasite genes and polymorphisms. Our lab was among the first to adapt CRISPR/Cas9 to engineer the T. gondii genome, and our plasmids are available to anyone through Addgene. We continue to improve these systems to enable genome-scale screening in parasites, which will allow exploration of the multitude of apicomplexan genes with unknown functions.
Sebastian Lourido Principal Investigator
Gabrielle McCauley Administrative Lab Manager
Emily Shortt Lab Manager
Faye Harling Technical Assistant
Sundeep Chakladar UROP
Kenneth Wei UROP
Mudita Goyal UROP
Tyler Smith Doctoral Student
Justin Roberts Doctoral Student
Alice Herneisen Doctoral Student
Alex Chan Doctoral Student
Chris Giuliano Doctoral Student
Michelle Peters Doctoral Student
Dominic Schwarz Doctoral Student
Dylan McCormick Doctoral Student
Dylan Valleau Postdoctoral Fellow
Human Frontier Science Program Fellow
Haley Licon Postdoctoral Fellow
Aditi Shukla Postdoctoral Fellow
Helen Hay Whitney Fellow
High-Resolution Mapping of Transcription Initiation in the Asexual Stages of Toxoplasma gondii
Markus BM, Waldman BS, Lorenzi HA, Lourido S
Front Cell Infect Microbiol. 2021 Jan 20;10:617998.
Identifying the Target of an Antiparasitic Compound in Toxoplasma Using Thermal Proteome Profiling.
Herneisen AL, Sidik SM, Markus BM, Drewry DH, Zuercher WJ, Lourido S.
ACS Chem Biol. 2020 Jul 17;15(7):1801-1807.
Functional and Computational Genomics Reveal Unprecedented Flexibility in Stage-Specific Toxoplasma Metabolism.
Krishnan A, Kloehn J, Lunghi M, Chiappino-Pepe A, Waldman BS, Nicolas D, Varesio E, Hehl A, Lourido S, Hatzimanikatis V, Soldati-Favre D.
Cell Host Microbe. 2020 Jan 27. pii: S1931-3128(20)30041-X.
Waldman BS, Schwarz D, Wadsworth II MS, Saeij JP, Shalek AK, Lourido S.
Cell. 2020 Jan 10. pii: S0092-8674(19)31375-3.
Plate-Based Quantification of Stimulated Toxoplasma Egress.
Shortt E, Lourido S.
Methods Mol Biol. 2020;2071:171-186.
High-Throughput Measurement of Microneme Secretion in Toxoplasma gondii.
Brown KM, Sibley LD, Lourido S.
Methods Mol Biol. 2020;2071:157-169.