The Cell Cycle Timing of Centromeric Chromatin Assembly in Drosophila Meiosis Is Distinct from Mitosis Yet Requires CAL1 and CENP-C
(A) Changes in the amount of CID at centromeres during meiosis II and in differentiating spermatids. Larval testes were fixed and stained with anti-CID antibody (green), and DNA is stained with DAPI (red). Scale bar: 1 µM. (B) Quantification of total centromeric CID fluorescence intensity per nucleus in meiosis II and in differentiating spermatids. Bars, standard errors. N = 799 total cells; 25, M7–M9; 114, M10–M11; 90, T1–T2; 295, T3; 196, T4–T5; 79, maturing spermatids. Note that Figures 2C and 4B are from the same experiment and to the same scale, normalized to the initial S1 intensity value. (C) Live imaging of GFP-CID (green) and H2Av-RFP (red) expression in M10–M11 (telophase II) and differentiating spermatids (T1–T3 and T4). Scale bar: 3 µM. (D) Quantification of total centromeric GFP-CID fluorescence intensity per nucleus from live imaging of stage M10–M11 (telophase II, n = 24) and T1–T3 (n = 36) and T4 (n = 38) spermatids. Bars, standard errors. (E) CID localization on spermatids in stage S1 primary spermatocytes and before (early canoe stage) and after (late canoe stage) protamine exchange in adult testes fixed and stained with anti-CID antibody (green), anti-histone (blue), and DAPI (red). Scale bar: 15 µM. (F) Quantification of total centromeric CID fluorescence intensity per nucleus S1 primary spermatocytes, and early and late canoe stage spermatids in adult testes. Bars, standard errors. Values for early and late canoe are scaled to the S1 value. N = 144 total cells; 52, S1; 25, early canoe; 67, late canoe. (G) CID localization in mature spermatozoa. Adult testes from GFP-CID (green) transgenic flies were fixed and stained with DAPI (red). Scale bar: 5 µM.