Figure 2.—

Telomere healing is abrogated in mre11-A470T cells. (A) Strains designed as described in materials and methods contain (telomere distal to proximal) the ADE2 marker, a telomere seed sequence (TSS), an HO cut site, subtelomeric sequences (SUBTEL), and telomeric sequences (TEL) (Diede and Gottschling 1999). In these cells, the HO-encoded endonuclease is under the control of a galactose-regulated HO endonuclease promoter (GAL-HO). (B) Southern blotting of SpeI-digested DNA using ADE2 as a probe was used to determine the cleavage of the recognition site and the addition of telomere repeats to the telomeric seed sequence. Southern blots were performed for strains that carried mre11-A470T, mre11Δ, or MRE11 alleles as shown on the top of the gel. M refers to the size marker lane. The image is derived from one gel that is spliced between the A470T-20 hr and mre11Δ-0 hr. The cells were synchronized in nocodazole and shifted to galactose for the time indicated on the bottom of the gel (in hr). Only MRE11 and mre11-A470T cells transformed with plasmid-borne MRE11 (data not shown) displayed full healing activity.