LI ET AL. SUPPLEMENTAL MATERIAL
ATF3
Ad-virus:
Infection (hrs): 0
4
8
16
24
ATF3
Actin
050207 PDX1 Real Time PCR
050207 PDX1 Real Time PCR
S. FIG. 1 MIN6 cells050207
were
infected
PDX1
Real Timewith
PCR Ad-tTA plus Ad-ATF3 for 0, 4, 8, 16 or 24h as
050207 PDX1 Real Time PCR
indicated, followed by immunoblot for ATF3 and actin.
1
Ad-β-Gal
Ad-BGal
Ad-ATF3
0
16
0
8
16
8
0.5
0.5
0.00
1
Ad-ATF3
0.5
168
8
0
8
16
16
16
Ad-BGal
050207 PDX1 Real Time PCR
Ad-β-Gal
Ad-BGal
Ad-ATF3
B
Relative PDX1 mRNA
0.5
Relative PDX1 mRNA
1.01
Relative PDX1 mRNA
A
0.5
050207 PDX1 Real Time
050207
PCRPDX1 Real Time PCR
1.01
Relative PDX1 mRNA
Ad-BGal
Relative Pdx1 mRNA
Relative
PDX1
mRNA
Relative
PDX1
mRNA
1
Ad-ATF3
Ad-BGal
Ad-BGal
Ad-BGal
Ad-ATF3
Ad-ATF3
Ad-ATF3
0.5
0.5
0.00
Ex-4
8
16
16
S. FIG. 2 The steady-state Pdx1 mRNA levels in the absence or presence of ATF3
expression. A: MIN6 cells were infected with Ad-tTA plus Ad-β-Gal or Ad-ATF3 for 8
or 16 h as indicated, followed by qRT-PCR for Pdx1. The standardized Pdx1 mRNA
level (against actin) from the vector control at each time point was arbitrarily defined as 1.
B: MIN6 cells infected as in A were cultured overnight in normal medium followed by
exendin-4 treatment (2.5 nM) in serum free medium for 8 h, and analyzed by qRT-PCR
for Pdx1. The standardized Pdx1 mRNA level from the vector control was arbitrarily
defined as 1.
1
S. TABLE 1. Primers used for RT-PCR/qRT-PCR and ChIP
Assay
Gene
Species
Forward primer (5’ to 3’)
Reverse primer (5’ to 3’)
RT-
ATF3
mouse/rat
GAGATGTCAGTCACCAAGTC
CAGTTTCTCTGACTCTTTCTGC
PCR
IRS2
mouse
GTGGGTTTCCAGAACGGCCT
ATGGGGCTGGTAGCGCTTCA
IRS2
rat
GTGGGTTTCCAGAACGGCCT
TGAGCAGCGTTGGTTGGA
Pdx1
mouse/rat
GAAATCCACCAAAGCTCACG
AATTCCTTCTCCAGCTCCAG
β-actin
mouse/rat
AAGATCAAGATCATTGCTCCTC
GGACTCATCGTACTCCTG
β-actin
rat
GGTAAAAAAATGCTGCACTG
GGTTTTATAGGACGCCAC
α-actin
rat
AATATGGCTTGGGAAGGG
TACCTGCTGCTCTGACTC
primer
mouse
GTGGGTTTCCAGAACGGCCT
CCCCCATCCTCAAGGTCAAA
set #1
rat
primer
mouse
CTATTACATCCAGAACAGGCG
ATGGCAGCTCGGTGCCTTTT
rat
TTACTCAGAAACACAGTCAC
CAACCTCAGATTCCTTTCAG
mouse/rat
TCTCCTATTACATCCAGAACAGG
TCATCAGAGCCATTCACTTGTC
ChIP
IRS2
GAGTTCTTGTCTCCTGGCTG
set# 2
primer
set #3
Primer
set #4
2