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. 2018 Jul 2;217(7):2383–2401. doi: 10.1083/jcb.201711097

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© 2018 Schiklenk et al.

This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).

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Figure 5. — Zas1-dependent control of Cnd1 or Peg1 expression is not solely responsible for condensation defects. (A) Comparison of protein levels by immunoblotting of whole-cell extracts from asynchronous cultures of strains derived from dissection of the same zas1⁺/zas1-Δaj3 tetrad (C4543, C4610, C4870, C4990) against the PK₆ tag fused to condensin subunits Cnd1, Cnd2, Cnd3, or Cut3. Immunoblotting against α-tubulin serves as loading control. (B) RT-qPCR signals for cnd1 and cnd2 transcripts in asynchronous zas1⁺ or zas1-Δaj3 mutant cells expressing cnd1 from either its endogenous promoter (cnd1⁺) or the cnd3 promoter (P_cnd3-cnd1). Graphs show mean ± SD of measurements from three independent tetrads (C4718). (C) Comparison of Cnd1 protein levels in asynchronous cultures (C4718) by immunoblotting with an antibody raised against the Cnd1 protein. Immunoblotting against α-tubulin serves as loading control. (D) Quantitation of the fraction of mitotic cells that display unequal chromosome segregation at 34°C of strains derived in E; n = 258 (zas1⁺, cnd1⁺), n = 231 (zas1⁺ P_cnd3-cnd1), n = 101 (zas1-Δaj3 cnd1⁺), and n = 114 (zas1-Δaj3 P_cnd3-cnd1) mitoses scored. (E) Chromosome condensation curves of FROS strains with the indicated genotypes derived by tetrad dissection of one parental diploid strain (C4984). (F) Tetrad analysis of a heterozygous zas1-Δaj3/zas1⁺ P_cnd3-cnd1/cnd1⁺ diploid fission yeast strain (C4718) imaged after 5 d at 25°C. (G) Tetrad analysis of heterozygous ΔSPBC887.16/SPBC887.16 and ΔSPBC713.13/SPBC713.13, ΔSPBC713.14c/SPBC713.14c diploid fission yeast strains (C4509, C4508) after 5 d at 25°C. Circles identify the position of spores bearing the ΔSPBC887.16 or ΔSPBC713.13, ΔSPBC713.14c genes, respectively. (H) Chromosome condensation curves of FROS strains with the indicated genotypes derived by tetrad dissection of one parental diploid strain (C5048). (I) Quantitation of the fraction of mitotic cells that display unequal chromosome segregation at 34°C of strains used in H; n = 89 zas1-Δaj3 P_cnd3-peg1 mitoses scored; other data from D.