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. Author manuscript; available in PMC: 2016 Feb 2.
Published in final edited form as: Curr Biol. 2015 Jan 22;25(3):348–356. doi: 10.1016/j.cub.2014.11.060

Figure 4. Cnp1 N-tail variants selectively reduce centromeric accumulation of the Cnp20/CENP-T branch of the CCAN.

Figure 4

A. Representative images of Cnp20-GFP and Cnp3-tomato in the indicated strains. Scale bar represents 5 μm. See also Fig. S4A.

B. Integrated fluorescence intensity of Cnp20-GFP, Mis6-GFP and Cnp3-tomato foci was measured and plotted for the indicate strains as in Fig. 1F&G. Error bars represent the standard deviation.

C. ChIP-qPCR with H3K9me2 antibody for the indicated conditions. The normalized ratio of products from Cnp1-enriched regions (cc) and heterochromatic outer repeats (otr) is displayed. See also Fig. S4B. Error bars represent the SD (n=3).

D. Serial dilutions of indicated strains harboring empty or Cnp20-overexpressing multicopy pREP1 plasmid were plated on minimal medium lacking leucine (PMG –LEU), with or without 15 μg/ml TBZ, and grown at 33°C. A ten-fold dilution series is shown for each strain.

E. Schematic summary of key findings. The Cnp1/CENP-A N-tail is required to recruit the Cnp20/CENP-T branch of the CCAN, which in turn is required for stable epigenetic propagation of centromeres (left). Cnp1 N-tail variants are normally loaded and support outer kinetochore assembly but the CENP-T branch of the CCAN is selectively reduced, increasing the probability of centromere inactivation and chromosome missegregation (right).