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Fig. 9. An AP-1-like element present in the 5'-flanking region of the 202 gene can bind to JunD/AP-1 transcription factor. A, left panel, subconfluent cultures of murine AKR-2B cells were incubated in growth medium supplemented with 10% FBS, and total cell extracts were prepared as described in "Materials and Methods." Samples containing equal amounts of proteins were analyzed by gel mobility shift assays using labeled oligonucleotide containing the AP-1 DNA-binding consensus sequence (probe; Santa Cruz Biotechnology). Extracts with labeled probe (Lane 2) or with labeled probe along with a 50-fold mole excess of cold oligonucleotide (Lane 3). As a control, probe without an extract was also run in Lane 1. Right panel, extracts were subjected to gel mobility shift assays using labeled probe containing the AP-1-like DNA-binding sequence present in the 202 gene promoter (Lanes 4–8). Probe without any extracts (Lane 4), with extracts along with cold oligonucleotide (containing the AP-1 consensus sequence) 2.5-fold mole excess (Lane 5), with 5-fold mole excess (Lane 6), with 10-fold mole excess (Lane 7), and with 20-fold mole excess (Lane 8). An arrow indicates the AP-1-specific band. B, under reduced serum conditions, increased levels of JunD are detected in the AP-1 DNA-binding complex. Subconfluent cultures of AKR-2B cells were incubated in growth medium supplemented with either 10% (Lanes 1–4) or 1% serum (Lanes 5 and 6) for 0 day (Lanes 1 and 2) or 2 days (Lanes 3–6). The nuclear extracts were prepared as described in "Materials and Methods," and samples containing equal amounts of proteins were analyzed by gel mobility shift assays using labeled oligonucleotide probe containing the AP-1 consensus sequence (same as that described in A above). The samples were incubated with anti-JunD antibody before subjecting to gel mobility shift assays (Lanes 2, 4, and 6) as described in "Materials and Methods." A thin arrow indicates the AP-1-specific band. A thick arrow indicates the JunD/AP-1 protein supershifted by the anti-JunD antibodies.





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HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Cancer Research Clinical Cancer Research
Cancer Epidemiology Biomarkers & Prevention Molecular Cancer Therapeutics
Molecular Cancer Research Cell Growth & Differentiation