Supplemental materials
Figure legends
Supplemental Figure 1. The XPF gene structure and conserved sequences. Three
tagSNPs were shown by arrows. Exons were boxed in red. Evolutionary conservation region
analysis in homo (H) and mouse (M) was carried out from the first intron to promoter region
with ClustalW 2.0 software.
Supplemental Figure 2. Effect of the XPF -357A>C polymorphism in promoter
transcriptional activity. (A) Schematic representation of reporter plasmids containing the
-357A or -357C allele, which was inserted upstream of the luciferase reporter gene in pGL3
basic plasmid. (B) The two constructs were transiently transfected into the NIH-3T3, HeLa,
and T24 cells, respectively. (C) The two constructs were transiently transfected into the
NIH-3T3 with or without HMGB2 siRNA and UV irradiation, respectively. The luciferase
activity of each construct was normalized against the internal control of Renilla luciferase.
The data indicate the mean values with the standard deviation (SD) from three independent
experiments. *, P < 0.05, compared with the construct counterpart.
Supplemental Figure 3. SDS-PAGE of the purified XPF -357A>C polymorphism
binding protein using the DNA-binding protein purification kit and MALDI-TOF MS
results. (A) Four purified proteins showed molecular masses of approximately from 32 to 66
KDa (indicated by arrow A, B, C and D). M, marker; E, eluted with 2.0 M KCl. (B) PMF of
the tryptic digestion of protein D. Using the MASCOT search engine, HMGB2 was identified
with a score of 202 and a sequence coverage of 52%. Peptides matched to the theoretical
masses were indicated in the picture. (C) Amino acid sequence of HMGB2. Peptides
identified by PMF were indicated by bold red.
Supplemental Figure 4. Association between the XPF -357A>C polymorphism and
mRNA expression. XPF transcript in bladder tumor tissues detected by real-time quantitative
RT-PCR. The frequency distributions of the AA, AC, and CC genotypes were 15, 5, and 2,
respectively. The fold change was normalized against -actin. *, P < 0.01 compared with the
AA genotypes.
Supplemental Figure 5. Immunohistochemical stainings of XPF in human bladder
tissues. A and B. Normal bladder tissues. C, D, E, and F. Bladder cancer tissues with different
genotypes of the XPF -357A>C polymorphism (C, D, AA genotype; E, AC genotype; F, CC
genotype). Decreased positive staining was observed in bladder cancer tissues with the
-357AC/CC genotypes, compared with the normal bladder tissues. Bladder cancer tissues
with the XPF -357AC/CC genotypes showed reduced staining compared with the -357AA
genotype. The staining in bladder tissue cells was mainly observed in the cytoplasm and
nucleus. Magnification, A and C  40, B, D, E, and F  400. Arrow, bladder mucosas.
Supplementary Table 1. Distribution of selected variables between the bladder cancer cases
and cancer-free control subjects
Variables
Age (meanSD)
Sex
Male
Female
Smoking status
Never
Ever
Former
Current
Pack-years smoked
0
0-20
>20
First-set (N=484)
Cases
Controls
(N=234)
(N=250)
N
%
N
%
Second-set (N=280)
Cases
Controls
(N=130)
(N=150)
N
%
N
%
63.812.4
63.510.9
63.012.1
63.510.9
192
42
82.0
18.0
194
56
77.6
22.4
106
24
81.5
18.5
112
38
74.7
25.3
101
133
62
71
43.2
56.8
26.5
30.3
152
98
30
68
60.8
39.2
12.0
27.2
59
71
34
37
45.4
54.6
26.1
28.5
92
58
16
42
61.3
38.7
10.7
28.0
101
73
60
43.2
31.2
25.6
152
50
48
60.8
20.0
19.2
59
39
32
45.4
30.0
24.6
92
28
30
61.3
18.7
20.0
Supplementary Table 2. Primary information of genotyped XPF SNPs in the first-set screening
Gene (accession
no.) and locus
NCBI
rs no.
Positiona
Location
Base change
XPF
(NM_005236)
16p13.3-p13.11
rs6498486
5236745
promoter
rs744154
5328160
rs31870
rs1799801
a
c
P for HWEc
SNP selection
0.233
0.403
-
0.197
0.272
0.079
tagSNP
0.244
0.184
0.196
0.808
tagSNP
0.211
0.254
0.248
0.117
tagSNP
HapMapb
Case
Control
A>C
0.233
0.308
intron 1
C>G
0.227
5350809
intron 9
G>A
5355037
exon 11
T>C
SNP position in NCBI dbSNP (http://www.ncbi.nlm.nih.gov/SNP).
b
MAF
MAF from the HapMap databases (http://www.hapmap.org).
HWE P value in the control group.
Supplemental Figure 1.
Supplemental Figure 2.
A
B
C
Supplemental Figure 3.
A
Intens. [a.u.]
B
8000
6000
4000
2000
0
800
1000
1200
1400
1600
m/z
C
Supplemental Figure 4.
Supplemental Figure 5.
A
B
C
D
E
F