FIG. 3.

Effects of TLC1 mutations on Rap1p binding in vitro. (A) Sequence of the conserved (8) consensus Rap1p binding site, which includes and extends the duplex GGTGT sequence that is seen to be bound directly in a Rap1p-DNA cocrystal structure (20). The Rap1 binding consensus sequence is aligned with a typical wild-type telomeric repeat, used as the labeled probe for gel shift analyses. (B) Gel shift analyses using ³²P-labeled wild-type telomeric repeat oligonucleotides, incubated with S100 whole cell extracts in the absence (lanes 1 and 10) or presence of 3,000, 1,000, 300, 100, 30, 10, 3, or 1 molar excess of unlabeled competitor oligonucleotides (lanes 2 through 9, respectively) and separated on a native gel. For all TLC1 alleles other than tlc1-476guA, only the shifted bands are shown.