{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,21]],"date-time":"2026-02-21T06:30:02Z","timestamp":1771655402971,"version":"3.50.1"},"reference-count":49,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2023,4,12]],"date-time":"2023-04-12T00:00:00Z","timestamp":1681257600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Key Research and Development Program of China","award":["2022YFD1800200"],"award-info":[{"award-number":["2022YFD1800200"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Protein prenylation is an important protein modification that is responsible for diverse physiological activities in eukaryotic cells. This modification is generally catalyzed by three types of prenyl transferases, which include farnesyl transferase (FT), geranylgeranyl transferase (GGT-1) and Rab geranylgeranyl transferase (GGT-2). Studies in malaria parasites showed that these parasites contain prenylated proteins, which are proposed to play multiple functions in parasites. However, the prenyl transferases have not been functionally characterized in parasites of subphylum Apicomplexa. Here, we functionally dissected functions of three of the prenyl transferases in the Apicomplexa model organism Toxoplasma gondii (T. gondii) using a plant auxin-inducible degron system. The homologous genes of the beta subunit of FT, GGT-1 and GGT-2 were endogenously tagged with AID at the C-terminus in the TIR1 parental line using a CRISPR-Cas9 approach. Upon depletion of these prenyl transferases, GGT-1 and GGT-2 had a strong defect on parasite replication. Fluorescent assay using diverse protein markers showed that the protein markers ROP5 and GRA7 were diffused in the parasites depleted with GGT-1 and GGT-2, while the mitochondrion was strongly affected in parasites depleted with GGT-1. Importantly, depletion of GGT-2 caused the stronger defect to the sorting of rhoptry protein and the parasite morphology. Furthermore, parasite motility was observed to be affected in parasites depleted with GGT-2. Taken together, this study functionally characterized the prenyl transferases, which contributed to an overall understanding of protein prenylation in T. gondii and potentially in other related parasites.<\/jats:p>","DOI":"10.3390\/ijms24087172","type":"journal-article","created":{"date-parts":[[2023,4,13]],"date-time":"2023-04-13T02:29:15Z","timestamp":1681352955000},"page":"7172","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["Prenyl Transferases Regulate Secretory Protein Sorting and Parasite Morphology in Toxoplasma gondii"],"prefix":"10.3390","volume":"24","author":[{"given":"Qiang-Qiang","family":"Wang","sequence":"first","affiliation":[{"name":"National Key Laboratory of Veterinary Public Health Security, School of Veterinary Medicine, China Agricultural University, Beijing 100193, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2252-4855","authenticated-orcid":false,"given":"Kai","family":"He","sequence":"additional","affiliation":[{"name":"National Key Laboratory of Veterinary Public Health Security, School of Veterinary Medicine, China Agricultural University, Beijing 100193, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Muhammad-Tahir","family":"Aleem","sequence":"additional","affiliation":[{"name":"Center for Gene Regulation in Health and Disease, Department of Biological, Geological and Environmental Sciences, College of Sciences and Health Professions, Cleveland State University, Cleveland, OH 44115, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shaojun","family":"Long","sequence":"additional","affiliation":[{"name":"National Key Laboratory of Veterinary Public Health Security, School of Veterinary Medicine, China Agricultural University, Beijing 100193, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2023,4,12]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"423","DOI":"10.1016\/j.ijpara.2003.12.009","article-title":"Toxoplasma gondii: The model apicomplexan","volume":"34","author":"Kim","year":"2004","journal-title":"Int. J. Parasitol."},{"key":"ref_2","first-page":"147","article-title":"Toxoplasmosis of the central nervous system in the acquired immunodeficiency syndrome","volume":"41","author":"Lanjewar","year":"1998","journal-title":"Indian J. Pathol. Microbiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"715","DOI":"10.1016\/j.parint.2018.07.004","article-title":"Congenital toxoplasmosis: An overview of the neurological and ocular manifestations","volume":"67","author":"Khan","year":"2018","journal-title":"Parasitol. Int."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"107","DOI":"10.1016\/S0950-3552(05)80149-X","article-title":"Toxoplasmosis in pregnancy","volume":"7","year":"1993","journal-title":"Bailliere\u2019s Clin. Obstet. Gynaecol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"2334","DOI":"10.1093\/jac\/dkx134","article-title":"Development of a murine vertical transmission model for Toxoplasma gondii oocyst infection and studies on the efficacy of bumped kinase inhibitor (BKI)-1294 and the naphthoquinone buparvaquone against congenital toxoplasmosis","volume":"72","author":"Muller","year":"2017","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"15936","DOI":"10.1073\/pnas.1208069109","article-title":"Endochin-like quinolones are highly efficacious against acute and latent experimental toxoplasmosis","volume":"109","author":"Doggett","year":"2012","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"54","DOI":"10.1016\/j.exppara.2014.12.007","article-title":"Triclosan and triclosan-loaded liposomal nanoparticles in the treatment of acute experimental toxoplasmosis","volume":"149","author":"Mossallam","year":"2015","journal-title":"Exp. Parasitol."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Mui, E.J., Schiehser, G.A., Milhous, W.K., Hsu, H., Roberts, C.W., Kirisits, M., Muench, S., Rice, D., Dubey, J.P., and Fowble, J.W. (2008). Novel triazine JPC-2067-B inhibits Toxoplasma gondii in vitro and in vivo. PLoS Negl. Trop. Dis., 2.","DOI":"10.1371\/journal.pntd.0000190"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"4843","DOI":"10.1128\/AAC.00462-10","article-title":"Activity of the histone deacetylase inhibitor FR235222 on Toxoplasma gondii: Inhibition of stage conversion of the parasite cyst form and study of new derivative compounds","volume":"54","author":"Maubon","year":"2010","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"647","DOI":"10.1128\/AAC.5.6.647","article-title":"Effect of clindamycin on acute and chronic toxoplasmosis in mice","volume":"5","author":"Araujo","year":"1974","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Li, Z.H., Ramakrishnan, S., Striepen, B., and Moreno, S.N. (2013). Toxoplasma gondii relies on both host and parasite isoprenoids and can be rendered sensitive to atorvastatin. PLoS Pathog., 9.","DOI":"10.1371\/journal.ppat.1003665"},{"key":"ref_12","first-page":"217","article-title":"Protein lipidation","volume":"283","author":"Kuhlmann","year":"2004","journal-title":"Methods Mol. Biol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"51","DOI":"10.1021\/cb500791f","article-title":"Protein prenylation: Enzymes, therapeutics, and biotechnology applications","volume":"10","author":"Palsuledesai","year":"2015","journal-title":"ACS Chem. Biol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"241","DOI":"10.1146\/annurev.bi.65.070196.001325","article-title":"Protein prenylation: Molecular mechanisms and functional consequences","volume":"65","author":"Zhang","year":"1996","journal-title":"Annu. Rev. Biochem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"8631","DOI":"10.1073\/pnas.88.19.8631","article-title":"Enzymatic modification of proteins with a geranylgeranyl isoprenoid","volume":"88","author":"Casey","year":"1991","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"4448","DOI":"10.1073\/pnas.88.10.4448","article-title":"Protein geranylgeranyltransferase of Saccharomyces cerevisiae is specific for Cys-Xaa-Xaa-Leu motif proteins and requires the CDC43 gene product but not the DPR1 gene product","volume":"88","author":"Finegold","year":"1991","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"7541","DOI":"10.1073\/pnas.87.19.7541","article-title":"Identification and preliminary characterization of protein-cysteine farnesyltransferase","volume":"87","author":"Manne","year":"1990","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"14603","DOI":"10.1016\/S0021-9258(18)98729-6","article-title":"Sequence dependence of protein isoprenylation","volume":"266","author":"Moores","year":"1991","journal-title":"J. Biol. Chem."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"81","DOI":"10.1016\/0092-8674(90)90242-7","article-title":"Inhibition of purified p21ras farnesyl:protein transferase by Cys-AAX tetrapeptides","volume":"62","author":"Reiss","year":"1990","journal-title":"Cell"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"14701","DOI":"10.1016\/S0021-9258(18)77164-0","article-title":"Polyisoprenylation of Ras in vitro by a farnesyl-protein transferase","volume":"265","author":"Schaber","year":"1990","journal-title":"J. Biol. Chem."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"429","DOI":"10.1016\/0092-8674(91)90460-G","article-title":"Protein farnesyltransferase and geranylgeranyltransferase share a common alpha subunit","volume":"65","author":"Seabra","year":"1991","journal-title":"Cell"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"5302","DOI":"10.1073\/pnas.88.12.5302","article-title":"A protein geranylgeranyltransferase from bovine brain: Implications for protein prenylation specificity","volume":"88","author":"Yokoyama","year":"1991","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1016\/S0898-6568(97)00120-4","article-title":"Membrane association and targeting of prenylated Ras-like GTPases","volume":"10","author":"Seabra","year":"1998","journal-title":"Cell Signal."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"48631","DOI":"10.1074\/jbc.M106470200","article-title":"Double prenylation by RabGGTase can proceed without dissociation of the mono-prenylated intermediate","volume":"276","author":"Niculae","year":"2001","journal-title":"J. Biol. Chem."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"16981","DOI":"10.1016\/S0021-9258(19)47327-4","article-title":"A novel prenyltransferase for a small GTP-binding protein having a C-terminal Cys-Ala-Cys structure","volume":"266","author":"Horiuchi","year":"1991","journal-title":"J. Biol. Chem."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"175","DOI":"10.1016\/S0166-6851(98)00065-6","article-title":"Protein prenyl transferase activities of Plasmodium falciparum","volume":"94","author":"Chakrabarti","year":"1998","journal-title":"Mol. Biochem. Parasitol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"42066","DOI":"10.1074\/jbc.M202860200","article-title":"Protein farnesyltransferase and protein prenylation in Plasmodium falciparum","volume":"277","author":"Chakrabarti","year":"2002","journal-title":"J. Biol. Chem."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"2553","DOI":"10.1128\/AAC.45.9.2553-2558.2001","article-title":"Limonene arrests parasite development and inhibits isoprenylation of proteins in Plasmodium falciparum","volume":"45","author":"Moura","year":"2001","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"761","DOI":"10.1016\/S0960-894X(01)00055-5","article-title":"Peptidomimetic inhibitors of protein farnesyltransferase show potent antimalarial activity","volume":"11","author":"Ohkanda","year":"2001","journal-title":"Bioorganic Med. Chem. Lett."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"e0205721","DOI":"10.1128\/mBio.02057-21","article-title":"Revisiting the Role of Toxoplasma gondii ERK7 in the Maintenance and Stability of the Apical Complex","volume":"12","author":"Tosetti","year":"2021","journal-title":"mBio"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Parker, K.E.R., Fairweather, S.J., Rajendran, E., Blume, M., McConville, M.J., Br\u00f6er, S., Kirk, K., and van Dooren, G.G. (2019). The tyrosine transporter of Toxoplasma gondii is a member of the newly defined apicomplexan amino acid transporter (ApiAT) family. PLoS Pathog., 15.","DOI":"10.1371\/journal.ppat.1007577"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"2236","DOI":"10.1038\/s41467-017-02341-2","article-title":"A conserved ankyrin repeat-containing protein regulates conoid stability, motility and cell invasion in Toxoplasma gondii","volume":"8","author":"Long","year":"2017","journal-title":"Nat. Commun."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"977","DOI":"10.1038\/s41467-023-36571-4","article-title":"The Toxoplasma micropore mediates endocytosis for selective nutrient salvage from host cell compartments","volume":"14","author":"Wan","year":"2023","journal-title":"Nat. Commun."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"e03622-20","DOI":"10.1128\/mBio.03622-20","article-title":"Identification and Molecular Dissection of IMC32, a Conserved Toxoplasma Inner Membrane Complex Protein That Is Essential for Parasite Replication","volume":"12","author":"Torres","year":"2021","journal-title":"mBio"},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Bansal, P., Antil, N., Kumar, M., Yamaryo-Botte, Y., Rawat, R.S., Pinto, S., Datta, K.K., Katris, N.J., Botte, C.Y., and Prasad, T.S.K. (2021). Protein kinase TgCDPK7 regulates vesicular trafficking and phospholipid synthesis in Toxoplasma gondii. PLoS Pathog., 17.","DOI":"10.1371\/journal.ppat.1009325"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"592","DOI":"10.1016\/j.copbio.2010.05.009","article-title":"How apicomplexan parasites move in and out of cells","volume":"21","author":"Sibley","year":"2010","journal-title":"Curr. Opin. Biotechnol."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Skillman, K.M., Diraviyam, K., Khan, A., Tang, K., Sept, D., and Sibley, L.D. (2011). Evolutionarily divergent, unstable filamentous actin is essential for gliding motility in apicomplexan parasites. PLoS Pathog., 7.","DOI":"10.1371\/journal.ppat.1002280"},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Egarter, S., Andenmatten, N., Jackson, A.J., Whitelaw, J.A., Pall, G., Black, J.A., Ferguson, D.J., Tardieux, I., Mogilner, A., and Meissner, M. (2014). The toxoplasma Acto-MyoA motor complex is important but not essential for gliding motility and host cell invasion. PLoS ONE, 9.","DOI":"10.1101\/001800"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"39","DOI":"10.1016\/j.molbiopara.2012.03.006","article-title":"Protein palmitoylation inhibition by 2-bromopalmitate alters gliding, host cell invasion and parasite morphology in Toxoplasma gondii","volume":"184","author":"Alonso","year":"2012","journal-title":"Mol. Biochem. Parasitol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1290","DOI":"10.1091\/mbc.e10-12-0939","article-title":"Actin depolymerizing factor controls actin turnover and gliding motility in Toxoplasma gondii","volume":"22","author":"Mehta","year":"2011","journal-title":"Mol. Biol. Cell"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Kremer, K., Kamin, D., Rittweger, E., Wilkes, J., Flammer, H., Mahler, S., Heng, J., Tonkin, C.J., Langsley, G., and Hell, S.W. (2013). An overexpression screen of Toxoplasma gondii Rab-GTPases reveals distinct transport routes to the micronemes. PLoS Pathog., 9.","DOI":"10.1371\/journal.ppat.1003213"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1157","DOI":"10.1111\/cmi.12426","article-title":"Toxoplasma gondii Vps11, a subunit of HOPS and CORVET tethering complexes, is essential for the biogenesis of secretory organelles","volume":"17","author":"Pastore","year":"2015","journal-title":"Cell Microbiol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"e00589-19","DOI":"10.1128\/mBio.00589-19","article-title":"Toxoplasma gondii Parasitophorous Vacuole Membrane-Associated Dense Granule Proteins Orchestrate Chronic Infection and GRA12 Underpins Resistance to Host Gamma Interferon","volume":"10","author":"Fox","year":"2019","journal-title":"mBio"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"e00851-19","DOI":"10.1128\/mSphere.00851-19","article-title":"Toxoplasma gondii Parasitophorous Vacuole Membrane-Associated Dense Granule Proteins Regulate Maturation of the Cyst Wall","volume":"5","author":"Guevara","year":"2020","journal-title":"mSphere"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"79","DOI":"10.1007\/978-1-4939-6472-7_6","article-title":"Development of CRISPR\/Cas9 for Efficient Genome Editing in Toxoplasma gondii","volume":"1498","author":"Shen","year":"2017","journal-title":"Methods Mol. Biol."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"e2728","DOI":"10.21769\/BioProtoc.2728","article-title":"Conditional Knockdown of Proteins Using Auxin-inducible Degron (AID) Fusions in Toxoplasma gondii","volume":"8","author":"Brown","year":"2018","journal-title":"Bio-Protocol"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"e0356121","DOI":"10.1128\/mbio.03561-21","article-title":"Novel CRK-Cyclin Complex Controls Spindle Assembly Checkpoint in Toxoplasma Endodyogeny","volume":"13","author":"Hawkins","year":"2021","journal-title":"mBio"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"4041","DOI":"10.1038\/s41467-019-11979-z","article-title":"A lipid-binding protein mediates rhoptry discharge and invasion in Plasmodium falciparum and Toxoplasma gondii parasites","volume":"10","author":"Suarez","year":"2019","journal-title":"Nat. Commun."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"e56635","DOI":"10.7554\/eLife.56635","article-title":"Essential function of the alveolin network in the subpellicular microtubules and conoid assembly in Toxoplasma gondii","volume":"9","author":"Tosetti","year":"2020","journal-title":"Elife"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/24\/8\/7172\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T19:15:03Z","timestamp":1760123703000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/24\/8\/7172"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,4,12]]},"references-count":49,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2023,4]]}},"alternative-id":["ijms24087172"],"URL":"https:\/\/doi.org\/10.3390\/ijms24087172","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,4,12]]}}}