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. 2022 Aug 27;13:5054. doi: 10.1038/s41467-022-32579-4

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Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — a Live-cell fluorescence imaging of stage II-V gametocytes transfected with the nuclear localisation signal (NLS)-mCherry reporter. NLS-mCherry (NLS-mCh, red) labels the elongated nucleus, with pointed extensions (yellow arrowheads). Subpellicular (green arrowheads) and nuclear (cyan arrowheads) populations of microtubules are labelled with Tubulin Tracker (TT, cyan). The nucleus contracts to a more spherical morphology and the nuclear and sub-pellicular microtubule populations are disassembled in stage V gametocytes. The chromatin (Hoechst, grayscale) is present as punctate features in the nucleus (white arrowheads). b Live-cell fluorescence imaging of stage II/III gametocytes in the PfEB1-GFP/NLS-mCherry co-transfectant parasite line. NLS-mCherry (NLS-mCh, red) delineates the nucleus; and PfEB1-GFP (EB1-GFP, green) is in the same compartment. Chromatin (Hoechst, grayscale) is distributed along the nuclear microtubules. (See Tubulin Tracker (TT, cyan) overlap with PfEB1-GFP, cyan arrows). Additional images are presented in Supplementary Fig. 3d. c Quantitative analysis of the contour lengths for the PfEB1-GFP (EB1) and nuclear microtubules (MT) labelled with Tubulin Tracker (mean ± SD is shown; n = 20 cells per stage). Differences in the length of nuclear microtubule features were evaluated by two-way ANOVA Tukey’s test, ****p < 0.0001. The blue dashed line indicates the limit of resolution (250 nm) of the microscope. d Images showing that the nuclear microtubules (as marked by anti-GFP (PfEB1-GFP), green) are depolymerised and the subpellicular microtubules (white arrowheads) are modified with polyglutamate (polyE, red) in stage IV gametocytes. Anti-β-tubulin (anti-β-tub, magenta) labels both nuclear and subpellicular microtubules. The chromatin was labelled with DAPI (grayscale). e Quantitative analysis of anti-polyE fluorescence intensity at different stages of development (mean ± SD is shown; n = 15 cells). Differences in polyE fluorescence signal were determined using a paired one-way ANOVA Tukey’s, I vs. II and II vs. IIIa: NS > 0.9999; IIIa vs. IIIb, IIIb vs. IV and IV vs. V: ****p < 0.0001. Differential interference contrast (DIC) images are shown. Scale bars: 2 μm. Additional images are presented in Supplementary Fig. 4d. Source data for Fig. 2c, e is provided in the Source Data file.