Characterization of epitope-tagged Rad9. (A) The relative
position of the ten histidine residues, single haemagglutinin (HA) epitope and
BRCA1 carboxy-terminal (BRCT) domains are indicated schematically. (B)
Rad9 western blot of extracts from exponentially growing strains that were
either mock treated (−) or irradiated with ultraviolet light (+).
(C) Exponentially growing cells of the strains indicated were serially
diluted onto YPD plates and irradiated with ultraviolet light at the doses
indicated. (D) Rad9 western blot of fractions from a Superose 6
gel-filtration column loaded with crude extracts from wild-type cells. Blots
from cells that were irradiated with ultraviolet light (+UV) or mock treated
(−UV) are indicated. (E) Rad9 western blot of fractions from a
20–35% glycerol sedimentation gradient loaded with crude cell extracts
from wild-type cells. Blots from cells that were irradiated with ultraviolet
light (+UV) and mock treated (−UV) are indicated. Hyperphosphorylated and
hypophosphorylated Rad9 and HH–Rad9 are indicated. The letters below the
panels in (D) and (E) indicate the positions of standard proteins
(masses, Stokes' radii and sedimentation coefficients are listed in the
supplementary information online).
A, aldolase; B, bovine serum albumin; Ch, chymotrypsinogen; DB, Dextran blue;
F, ferritin; O, ovalbumin; T, thyroglobulin; WT, wild-type.