{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2023,12,26]],"date-time":"2023-12-26T11:17:14Z","timestamp":1703589434141},"reference-count":70,"publisher":"Wiley","issue":"7","license":[{"start":{"date-parts":[[2018,8,17]],"date-time":"2018-08-17T00:00:00Z","timestamp":1534464000000},"content-version":"vor","delay-in-days":47,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"name":"Programa Estrat\u00e9gico de Apoio \u00e0 Pesquisa em Sa\u00fade (PAPES\/FIOCRUZ, 538 Brazil)","award":["407775\/2012-9-CNPq CAPES"],"award-info":[{"award-number":["407775\/2012-9-CNPq CAPES"]}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Cytometry Pt A"],"published-print":{"date-parts":[[2018,7]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Trypanosomatid parasites are causative agents of neglected human diseases. Their lineage diverged early from the common eukaryotic ancestor, and they evolved singular mechanisms of gene expression that are crucial for their survival. Studies on unusual and essential molecular pathways lead to new drug targets. In this respect, assays to analyze transcriptional activity will provide useful information to identify essential and specific factors. However, the current methods are laborious and do not provide global and accurate measures. For this purpose, a previously reported radiolabeling in vitro nascent mRNA methodology was used to establish an alternative fluorescent\u2010based assay that is able to precisely quantify nascent mRNA using both flow cytometry and a high\u2010content image system. The method allowed accurate and global measurements in <jats:italic>Trypanosoma brucei<\/jats:italic>, a representative species of trypanosomatid parasites. We obtained data demonstrating that approximately 70% of parasites from a population under normal growth conditions displayed mRNA transcriptional activity, whilst the treatment with \u03b1\u2010amanitin (75 \u00b5g\/ml) inhibited the polymerase II activity. The adaptation of the method also allowed the analyses of the transcriptional activity during the cell cycle. Therefore, the methodology described herein contributes to obtaining precise measurements of transcriptional rates using multiparametric analysis. This alternative method can facilitate investigations of genetic and biochemical processes in trypanosome parasites and consequently provide additional information related to new treatment or prophylaxis strategies involving these important human parasites.<\/jats:p>","DOI":"10.1002\/cyto.a.23387","type":"journal-article","created":{"date-parts":[[2018,8,17]],"date-time":"2018-08-17T22:31:42Z","timestamp":1534545102000},"page":"727-736","update-policy":"http:\/\/dx.doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Fluorescence\u2010based assay for accurate measurement of transcriptional activity in trypanosomatid parasites"],"prefix":"10.1002","volume":"93","author":[{"given":"Priscila M.","family":"Hiraiwa","sequence":"first","affiliation":[{"name":"Flow Cytometry Facility, Instituto Carlos Chagas, FIOCRUZ  Paran\u00e1 Brazil"},{"name":"Laborat\u00f3rio de Regula\u00e7\u00e3o da Express\u00e3o G\u00eanica, Instituto Carlos Chagas, FIOCRUZ  Paran\u00e1 Brazil"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alessandra M.","family":"de Aguiar","sequence":"additional","affiliation":[{"name":"Flow Cytometry Facility, Instituto Carlos Chagas, FIOCRUZ  Paran\u00e1 Brazil"},{"name":"Laborat\u00f3rio de Biologia B\u00e1sica de C\u00e9lulas\u2010Tronco, Instituto Carlos Chagas, FIOCRUZ  Paran\u00e1 Brazil"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Andr\u00e9a R.","family":"\u00c1vila","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Regula\u00e7\u00e3o da Express\u00e3o G\u00eanica, Instituto Carlos Chagas, FIOCRUZ  Paran\u00e1 Brazil"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2018,8,17]]},"reference":[{"key":"e_1_2_6_2_1","first-page":"1","article-title":"Research priorities for Chagas disease, human African trypanosomiasis and leishmaniasis","author":"World Health Organization","year":"2012","journal-title":"World Health Org Tech Rep Ser"},{"key":"e_1_2_6_3_1","doi-asserted-by":"publisher","DOI":"10.1038\/nrmicro.2016.193"},{"key":"e_1_2_6_4_1","doi-asserted-by":"publisher","DOI":"10.1042\/bse0510001"},{"key":"e_1_2_6_5_1","doi-asserted-by":"publisher","DOI":"10.1093\/emboj\/21.8.1881"},{"key":"e_1_2_6_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.mib.2007.10.001"},{"key":"e_1_2_6_7_1","doi-asserted-by":"publisher","DOI":"10.1155\/2010\/525241"},{"key":"e_1_2_6_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.molbiopara.2005.12.008"},{"key":"e_1_2_6_9_1","doi-asserted-by":"publisher","DOI":"10.1128\/EC.2.5.830-840.2003"},{"key":"e_1_2_6_10_1","doi-asserted-by":"publisher","DOI":"10.1016\/0092-8674(90)90674-4"},{"key":"e_1_2_6_11_1","doi-asserted-by":"publisher","DOI":"10.1042\/bst0280527"},{"key":"e_1_2_6_12_1","doi-asserted-by":"publisher","DOI":"10.1128\/EC.3.2.506-517.2004"},{"key":"e_1_2_6_13_1","doi-asserted-by":"publisher","DOI":"10.1101\/gad.1790409"},{"key":"e_1_2_6_14_1","doi-asserted-by":"crossref","unstructured":"ThomasS GreenA SturmNR CampbellDA MylerPJ.Histone acetylations mark origins of polycistronic transcription inLeishmania major. 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