Nucleolar Division in the Promastigote Stage of Leishmania major Parasite: A Nop56 Point of View

Figure 2

Nop56 expression in trypanosomatids and subcellular localization in L. major. (a) Western blot analysis of protein extracts from L. major promastigotes (Lm), T. brucei procyclic forms (Tb), and T. cruzi epimastigotes (Tc) was performed using an LmNop56 polyclonal antibody. (b) Indirect immunofluorescence experiment conducted with the same anti-LmNop56 serum and an anti-mouse IgG antibody conjugated with Alexa Fluor® 488 dye. Nuclei (N) and kinetoplast (K) in L. major promastigotes were counterstained with propidium iodide. Nucleolar (No) localization of LmNop56 was analyzed in single optical sections obtained by confocal microscopy. (c) Double indirect immunofluorescence assay carried out with antibodies raised against histone H4 and LmNop56. Histone H4 was revealed with anti-rabbit IgG coupled with Alexa Fluor® 488 (green), and LmNop56 with anti-mouse IgG conjugated with Alexa Fluor® 568 (red). (d) Double indirect immunofluorescence experiment performed with transgenic promastigotes expressing Elp3b-PTP. The antibodies employed were anti-LmNop56 and anti-Prot C. LmNop56 (red) was detected as indicated in panel (c), whereas Elp3b was revealed with anti-rabbit IgG coupled with Alexa Fluor® 488 (green). Images shown in panels (c) and (d) were obtained with an epifluorescence microscope; in these same panels, DNA was stained with DAPI (blue). Size bars represent 2 μm.

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