{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,5,7]],"date-time":"2026-05-07T00:23:00Z","timestamp":1778113380662,"version":"3.51.4"},"posted":{"date-parts":[[2024,2,1]]},"group-title":"Microbiology","reference-count":76,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2024,2,1]],"date-time":"2024-02-01T00:00:00Z","timestamp":1706745600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2024,10,21]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                <jats:p>\n                  <jats:italic>Plasmodium<\/jats:italic>\n                  malaria parasites retain an essential mitochondrional electron transport chain (ETC) that is critical for growth within humans and mosquitoes and a key antimalarial drug target. ETC function requires cytochromes\n                  <jats:italic>c<\/jats:italic>\n                  and\n                  <jats:italic>c<\/jats:italic>\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  that are unusual among heme proteins due to their covalent binding to heme via conserved CXXCH sequence motifs. Heme attachment to these proteins in most eukaryotes requires the mitochondrial enzyme holocytochrome\n                  <jats:italic>c<\/jats:italic>\n                  synthase (HCCS) that binds heme and the apo cytochrome to facilitate biogenesis of the mature cytochrome\n                  <jats:italic>c<\/jats:italic>\n                  or\n                  <jats:italic>c<\/jats:italic>\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  . Although humans encode a single bifunctional HCCS that attaches heme to both proteins,\n                  <jats:italic>Plasmodium<\/jats:italic>\n                  parasites are like yeast and encode two separate HCCS homologs thought to be specific for heme attachment to cyt\n                  <jats:italic>c<\/jats:italic>\n                  (HCCS) or cyt\n                  <jats:italic>c<\/jats:italic>\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  (HCC\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  S). To test the function and specificity of\n                  <jats:italic>P. falciparum<\/jats:italic>\n                  HCCS and HCC\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  S, we used CRISPR\/Cas9 to tag both genes for conditional expression. HCC\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  S knockdown selectively impaired cyt\n                  <jats:italic>c<\/jats:italic>\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  biogenesis and caused lethal ETC dysfunction that was not reversed by over-expression of HCCS. Knockdown of HCCS caused a more modest growth defect but strongly sensitized parasites to mitochondrial depolarization by proguanil, revealing key defects in ETC function. These results and prior heterologous studies in\n                  <jats:italic>E. coli<\/jats:italic>\n                  of cyt\n                  <jats:italic>c<\/jats:italic>\n                  hemylation by\n                  <jats:italic>P. falciparum<\/jats:italic>\n                  HCCS and HCC\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  S strongly suggest that both homologs are essential for mitochondrial ETC function and have distinct specificities for biogenesis of cyt\n                  <jats:italic>c<\/jats:italic>\n                  and\n                  <jats:italic>c<\/jats:italic>\n                  <jats:bold>\n                    <jats:sub>1<\/jats:sub>\n                  <\/jats:bold>\n                  , respectively, in parasites. This study lays a foundation to develop novel strategies to selectively block ETC function in malaria parasites.\n                <\/jats:p>","DOI":"10.1101\/2024.02.01.575742","type":"posted-content","created":{"date-parts":[[2024,2,1]],"date-time":"2024-02-01T23:50:15Z","timestamp":1706831415000},"source":"Crossref","is-referenced-by-count":9,"title":["Biogenesis of cytochromes\n                  <i>c<\/i>\n                  and\n                  <i>c<\/i>\n                  <sub>1<\/sub>\n                  in the electron transport chain of malaria parasites"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-6751-8304","authenticated-orcid":false,"given":"Aldo E.","family":"Garc\u00eda-Guerrero","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rebecca G.","family":"Marvin","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7473-5822","authenticated-orcid":false,"given":"Amanda Mixon","family":"Blackwell","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3464-3042","authenticated-orcid":false,"given":"Paul A.","family":"Sigala","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2024102404201126000_2024.02.01.575742v3.1","unstructured":"WHO. World Malaria Report; Geneva, Switzerland, 2023."},{"key":"2024102404201126000_2024.02.01.575742v3.2","doi-asserted-by":"publisher","DOI":"10.2450\/2017.0293-16"},{"key":"2024102404201126000_2024.02.01.575742v3.3","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-micro-091313-103537"},{"key":"2024102404201126000_2024.02.01.575742v3.4","doi-asserted-by":"publisher","DOI":"10.1089\/ars.2012.4539"},{"key":"2024102404201126000_2024.02.01.575742v3.5","doi-asserted-by":"publisher","DOI":"10.1111\/febs.15445"},{"key":"2024102404201126000_2024.02.01.575742v3.6","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.micro.51.1.97"},{"key":"2024102404201126000_2024.02.01.575742v3.7","doi-asserted-by":"publisher","DOI":"10.1038\/nm.4381"},{"key":"2024102404201126000_2024.02.01.575742v3.8","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pmed.1001169"},{"key":"2024102404201126000_2024.02.01.575742v3.9","doi-asserted-by":"publisher","DOI":"10.1016\/0006-2952(92)90213-3"},{"key":"2024102404201126000_2024.02.01.575742v3.10","doi-asserted-by":"publisher","DOI":"10.1038\/nature05572"},{"key":"2024102404201126000_2024.02.01.575742v3.11","doi-asserted-by":"publisher","DOI":"10.1126\/science.aat9446"},{"key":"2024102404201126000_2024.02.01.575742v3.12","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-019-0973-1"},{"key":"2024102404201126000_2024.02.01.575742v3.13","doi-asserted-by":"publisher","DOI":"10.1126\/scitranslmed.3005029"},{"key":"2024102404201126000_2024.02.01.575742v3.14","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-019-55506-y"},{"key":"2024102404201126000_2024.02.01.575742v3.15","doi-asserted-by":"publisher","DOI":"10.1128\/EC.05049-11"},{"key":"2024102404201126000_2024.02.01.575742v3.16","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.71636"},{"key":"2024102404201126000_2024.02.01.575742v3.17","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2301047120"},{"key":"2024102404201126000_2024.02.01.575742v3.18","first-page":"166","article-title":"Cyclization-blocked proguanil as a strategy to improve the antimalarial activity of atovaquone. Commun","volume":"2","year":"2019","journal-title":"Biol"},{"key":"2024102404201126000_2024.02.01.575742v3.19","doi-asserted-by":"publisher","DOI":"10.1016\/j.tibs.2015.05.006"},{"key":"2024102404201126000_2024.02.01.575742v3.20","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbamcr.2008.06.017"},{"key":"2024102404201126000_2024.02.01.575742v3.21","doi-asserted-by":"publisher","DOI":"10.1021\/ar030266l"},{"key":"2024102404201126000_2024.02.01.575742v3.22","doi-asserted-by":"publisher","DOI":"10.1093\/molbev\/msv021"},{"key":"2024102404201126000_2024.02.01.575742v3.23","doi-asserted-by":"publisher","DOI":"10.1002\/j.1460-2075.1987.tb04744.x"},{"key":"2024102404201126000_2024.02.01.575742v3.24","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1213897109"},{"key":"2024102404201126000_2024.02.01.575742v3.25","doi-asserted-by":"crossref","unstructured":"Sutherland, M. C. ; Mendez, D. L. ; Babbitt, S. E. ; Tillman, D. E. ; Melnikov, O. ; Tran, N. L. ; Prizant, N. T. ; Collier, A. L. ; Kranz, R. G . In vitro reconstitution reveals major differences between human and bacterial cytochrome c synthases. Elife 2021, 10.","DOI":"10.7554\/eLife.64891"},{"key":"2024102404201126000_2024.02.01.575742v3.26","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M308881200"},{"key":"2024102404201126000_2024.02.01.575742v3.27","doi-asserted-by":"publisher","DOI":"10.1111\/j.1432-1033.1992.tb17146.x"},{"key":"2024102404201126000_2024.02.01.575742v3.28","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.RA119.011605"},{"key":"2024102404201126000_2024.02.01.575742v3.29","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2017.06.030"},{"key":"2024102404201126000_2024.02.01.575742v3.30","doi-asserted-by":"publisher","DOI":"10.1126\/science.aap7847"},{"key":"2024102404201126000_2024.02.01.575742v3.31","doi-asserted-by":"publisher","DOI":"10.1016\/j.molbiopara.2016.08.003"},{"key":"2024102404201126000_2024.02.01.575742v3.32","doi-asserted-by":"publisher","DOI":"10.1021\/bi500704p"},{"key":"2024102404201126000_2024.02.01.575742v3.33","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1615929114"},{"key":"2024102404201126000_2024.02.01.575742v3.34","doi-asserted-by":"crossref","unstructured":"Abramson, J. ; Adler, J. ; Dunger, J. ; Evans, R. ; Green, T. ; Pritzel, A. ; Ronneberger, O. ; Willmore, L. ; Ballard, A. J. ; Bambrick, J. ; Bodenstein, S. W. ; Evans, D. A. ; Hung, C. C. ; O\u2019Neill, M. ; Reiman, D. ; Tunyasuvunakool, K. ; Wu, Z. ; Zemgulyte, A. ; Arvaniti, E. ; Beattie, C. ; Bertolli, O. ; Bridgland, A. ; Cherepanov, A. ; Congreve, M. ; Cowen-Rivers, A. I. ; Cowie, A. ; Figurnov, M. ; Fuchs, F. B. ; Gladman, H. ; Jain, R. ; Khan, Y. A. ; Low, C. M. R. ; Perlin, K. ; Potapenko, A. ; Savy, P. ; Singh, S. ; Stecula, A. ; Thillaisundaram, A. ; Tong, C. ; Yakneen, S. ; Zhong, E. D. ; Zielinski, M. ; Zidek, A. ; Bapst, V. ; Kohli, P. ; Jaderberg, M. ; Hassabis, D. ; Jumper, J. M . Accurate structure prediction of biomolecular interactions with AlphaFold 3. Nature 2024.","DOI":"10.1038\/s41586-024-07487-w"},{"key":"2024102404201126000_2024.02.01.575742v3.35","doi-asserted-by":"publisher","DOI":"10.1016\/j.febslet.2014.07.026"},{"key":"2024102404201126000_2024.02.01.575742v3.36","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M114.593509"},{"key":"2024102404201126000_2024.02.01.575742v3.37","doi-asserted-by":"crossref","unstructured":"Ganesan, S. M. ; Falla, A. ; Goldfless, S. J. ; Nasamu, A. S. ; Niles, J. C . Synthetic RNA-protein modules integrated with native translation mechanisms to control gene expression in malaria parasites. Nat. Commun. 2016, 7, 10727.","DOI":"10.1038\/ncomms10727"},{"key":"2024102404201126000_2024.02.01.575742v3.38","doi-asserted-by":"crossref","unstructured":"Maruthi, M. ; Ling, L. ; Zhou, J. ; Ke, H . Dispensable Role of Mitochondrial Fission Protein 1 (Fis1) in the Erythrocytic Development of Plasmodium falciparum. mSphere 2020, 5.","DOI":"10.1128\/mSphere.00579-20"},{"key":"2024102404201126000_2024.02.01.575742v3.39","doi-asserted-by":"crossref","unstructured":"Okada, M. ; Rajaram, K. ; Swift, R. P. ; Mixon, A. ; Maschek, J. A. ; Prigge, S. T. ; Sigala, P. A . Critical role for isoprenoids in apicoplast biogenesis by malaria parasites. Elife 2022, 11, eLife.73208.","DOI":"10.7554\/eLife.73208"},{"key":"2024102404201126000_2024.02.01.575742v3.40","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(18)81780-X"},{"key":"2024102404201126000_2024.02.01.575742v3.41","doi-asserted-by":"publisher","DOI":"10.1002\/pmic.201400617"},{"key":"2024102404201126000_2024.02.01.575742v3.42","doi-asserted-by":"publisher","DOI":"10.1002\/biot.201100155"},{"key":"2024102404201126000_2024.02.01.575742v3.43","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2014.00172"},{"key":"2024102404201126000_2024.02.01.575742v3.44","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-021-23919-x"},{"key":"2024102404201126000_2024.02.01.575742v3.45","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1009301"},{"key":"2024102404201126000_2024.02.01.575742v3.46","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1009211"},{"key":"2024102404201126000_2024.02.01.575742v3.47","doi-asserted-by":"crossref","unstructured":"Sheokand, P. K. ; Pradhan, S. ; Maclean, A. E. ; Muhleip, A. ; Sheiner, L . Plasmodium falciparum Mitochondrial Complex III, the Target of Atovaquone, Is Essential for Progression to the Transmissible Sexual Stages. Int J Mol Sci 2024, 25.","DOI":"10.1101\/2024.01.09.574740"},{"key":"2024102404201126000_2024.02.01.575742v3.48","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-023-42030-x"},{"key":"2024102404201126000_2024.02.01.575742v3.49","doi-asserted-by":"publisher","DOI":"10.1126\/science.aad9279"},{"key":"2024102404201126000_2024.02.01.575742v3.50","doi-asserted-by":"publisher","DOI":"10.1038\/33612"},{"key":"2024102404201126000_2024.02.01.575742v3.51","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(18)68224-9"},{"key":"2024102404201126000_2024.02.01.575742v3.52","doi-asserted-by":"crossref","unstructured":"Muhleip, A. ; Kock Flygaard, R. ; Ovciarikova, J. ; Lacombe, A. ; Fernandes, P. ; Sheiner, L. ; Amunts, A . ATP synthase hexamer assemblies shape cristae of Toxoplasma mitochondria. Nat. Commun. 2021, 12, 120.","DOI":"10.1038\/s41467-020-20381-z"},{"key":"2024102404201126000_2024.02.01.575742v3.53","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2019.07.019"},{"key":"2024102404201126000_2024.02.01.575742v3.54","doi-asserted-by":"publisher","DOI":"10.1128\/MMBR.00001-09"},{"key":"2024102404201126000_2024.02.01.575742v3.55","doi-asserted-by":"publisher","DOI":"10.1111\/j.1742-4658.2008.06380.x"},{"key":"2024102404201126000_2024.02.01.575742v3.56","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkq366"},{"key":"2024102404201126000_2024.02.01.575742v3.57","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M114.615831"},{"key":"2024102404201126000_2024.02.01.575742v3.58","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1003522"},{"key":"2024102404201126000_2024.02.01.575742v3.59","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.00148-16"},{"key":"2024102404201126000_2024.02.01.575742v3.60","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1006511"},{"key":"2024102404201126000_2024.02.01.575742v3.61","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1006396"},{"key":"2024102404201126000_2024.02.01.575742v3.62","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.09143"},{"key":"2024102404201126000_2024.02.01.575742v3.63","doi-asserted-by":"publisher","DOI":"10.1002\/j.1460-2075.1992.tb05074.x"},{"key":"2024102404201126000_2024.02.01.575742v3.64","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0135896"},{"key":"2024102404201126000_2024.02.01.575742v3.65","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymgme.2019.01.006"},{"key":"2024102404201126000_2024.02.01.575742v3.66","doi-asserted-by":"crossref","unstructured":"Spillman, N. J. ; Beck, J. R. ; Ganesan, S. M. ; Niles, J. C. ; Goldberg, D. E . The chaperonin TRiC forms an oligomeric complex in the malaria parasite cytosol. Cell Microbiol. 2017, 19, cmi.12719.","DOI":"10.1111\/cmi.12719"},{"key":"2024102404201126000_2024.02.01.575742v3.67","doi-asserted-by":"publisher","DOI":"10.1038\/345253a0"},{"key":"2024102404201126000_2024.02.01.575742v3.68","first-page":"50","article-title":"Chloroquine sensitivity of isolates of Plasmodium falciparum adapted to in vitro culture","volume":"33","year":"1981","journal-title":"Trop. Geogr. Med"},{"key":"2024102404201126000_2024.02.01.575742v3.69","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/20.11.2902"},{"key":"2024102404201126000_2024.02.01.575742v3.70","doi-asserted-by":"publisher","DOI":"10.1016\/0304-3894(92)87011-4"},{"key":"2024102404201126000_2024.02.01.575742v3.71","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/25.17.3389"},{"key":"2024102404201126000_2024.02.01.575742v3.72","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-62703-646-7_6"},{"key":"2024102404201126000_2024.02.01.575742v3.73","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btp033"},{"key":"2024102404201126000_2024.02.01.575742v3.74","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkw256"},{"key":"2024102404201126000_2024.02.01.575742v3.75","doi-asserted-by":"publisher","DOI":"10.1038\/S41586-021-03819-2"},{"key":"2024102404201126000_2024.02.01.575742v3.76","unstructured":"MacPyMOL: A PyMOL-based Molecular Graphics Application for MacOS X; DeLano Scientific LLC: Palo Alto, CA, 2007. http:\/\/www.pymol.org (accessed."}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2024.02.01.575742","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T05:53:10Z","timestamp":1768456390000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2024.02.01.575742"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,2,1]]},"references-count":76,"URL":"https:\/\/doi.org\/10.1101\/2024.02.01.575742","relation":{"is-preprint-of":[{"id-type":"doi","id":"10.1021\/acsinfecdis.4c00450","asserted-by":"subject"}]},"subject":[],"published":{"date-parts":[[2024,2,1]]},"subtype":"preprint"}}