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Supplementary Figure Legends
Supplementary Figure 1: Densitometry traces of MNase ladders at MAT.
Lanes chosen for analysis from the Southern blots shown in Figures 1-3 show equivalent patterns of
MNase digestion (~7-8 oligosomes) in bulk chromatin (Supplemental Figure 2).
Supplementary Figure 2: MNase digestion of bulk chromatin.
Nuclei were prepared from wild-type, mre11, and arp8 strains before and at 60 min intervals after
DSB induction, and MNase digestion was performed as described in Figure 1. Following agarose gel
electrophoresis of extracted DNA, gels were stained with ethidium bromide and photographed using an
Ultra-Lum gel documentation system.
Supplementary Figure 3: -H2A levels and histone eviction profiles in mre11∆ and arp8∆ mutants.
(a) ChIP was performed in wild-type or arp8∆ after DSB induction using -H2A antibodies. DNA was
analyzed by real time PCR on the right side of the DSB. (b) ChIP was performed with anti-Flag
antibodies in arp8 expressing Flag-H2B or Flag-H3 at times up to 180 min after DSB formation.
Analysis of DNA ~6 Kb on either side of the DSB (“0”) was determined by real time PCR. For FlagH3, the 180 min PCR results have been superimposed on the 0-120 min PCR data shown in Figure 3a.
(c) ChIP was performed in wild-type or mre11 after DSB induction using -H2A antibodies. DNA
was analyzed by real time PCR on the right side of the DSB. Data are means +/- s.e.m. 
Supplementary Figure 4: MAT DNA is cleaved and resected in an arp8∆ mutant.
(a) After DSB induction at MAT, DNA was extracted from wild-type and arp8∆ and digested with
StyI. Loss of 1.9 Kb (left) and 1.8 Kb (right) StyI fragments correlates with MAT DNA cleavage. Loss
of 1.4 Kb fragment (left) and 2.2 Kb and 0.7 Kb fragments (right), and gain of fragments larger than 1.9
and 2.2 Kb, correlate with 5’ to 3’ DNA resection. Top: Southern blot hybridization of StyI cleaved
DNA using DNA probes that hybridize to the left or right side of the DSB. The bracket represents
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single-stranded DNA resulting from resection. Bottom: Quantitation of 1.4 Kb (left), 2.2 Kb and 0.7
Kb (right) StyI fragments by phosphoimager analysis; all values were normalized to the level of ACT1
DNA. (b) After induction of a DSB at MAT DNA samples were taken from wild-type and arp8∆ for
Southern blot analysis (left) or ChIP (right). The percent MAT DNA cleaved was calculated by analysis
of the loss of 1.9 Kb and 1.8 Kb StyI fragments on the left and right side of the DSB, respectively, and
by qPCR in real time, using primers that span the DSB. (c) Single-stranded DNA turnover was
measured by quantitating two bands shown in brackets in the right panel of (a). Data are represented as
the fraction of maximum signal as described1.
Supplementary Figure 5: Role of -H2A in recruitment of Ino80 to the MAT DSB.
(a) ChIP of Ino80-Myc was performed in wildtype and hta1/hta2-S129* strains after induction of a
DSB at MAT. (b) ChIP of Ino80-Myc was performed in the same strains before induction of the
MAT DSB. For reference, the level of Ino80-Myc associated with the transcriptionally active POL5
gene is 1.5-2 fold higher than the level of Ino80-Myc at TELV, a nontranscribed control region. Data
are means +/- s.e.m.
Supplementary Figure 6: DNA repair phenotypes of an arp8∆ mutant.
(a) Plasmid rejoining assay. A wild type strain and an arp8∆ or mre11∆ mutant were transformed with
uncut or HindIII digested plasmid pRS316, and selection was performed on Ura- medium. The data
represent the ratio of Ura+ transformants from cut pRS316/uncut pRS316 obtained from 4 independent
experiments. (b) Sensitivity to an unrepairable DSB. A
strain containing GAL-
HO and rad52∆, arp8∆, or hta1/hta2-S129* mutations was induced with galactose for 2 hrs to create a
DSB at MAT. Glucose was added to stop DNA cleavage, and cells were plated in triplicate onto YPD
plates. Colonies were counted after 2-3 days incubation at 30° and the results are expressed as %
survival relative to uninduced strains. (c) Sensitivity to DNA damaging agents. Wild type, mre11∆,
arp8∆, rad52∆, arp8∆ rad52∆, and hta1/hta2-S129* strains were grown in YPD to O.D.600=0.6 and 102
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fold serial dilutions were spotted onto YPD, YPD + 25 mM HU, and YPD + 0.02% MMS. Cells were
incubated at 30° for 2-3 days before photographing.
Reference
1. Llorente, B. & Symington, L. S. The Mre11 nuclease is not required for 5' to 3' resection at multiple
HO induced double-strand breaks. Mol. Cell. Biol. 24, 9682-9694 (2004).
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