PMC:1920263 / 32355-33538 JSONTXT

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6. 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PCR primer pairs specific for the A3G promoter (upper panel) or the A3G gene (lower panel) were used. As positive controls, the sheared and cross-linked DNA before the immunoprecipitation step (input) or a plasmid carrying the target sequence (plasmid) was used as template."}

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6. Sp1 and Sp3 bind to the GC-box present in the A3G promoter. (A) Nuclear extracts of A3.01 T cells were incubated with a 32P-labeled commercial Sp1 oligonucleotide probe (Sp1cons) or labeled probes homologous to the unmodified or mutated E2 region (see Figure 1) of the A3G promoter (APO-Sp1/3 and APO-Sp1/3mut). Protein–DNA complexes were separated by polyacrylamide electrophoresis and detected by autoradiography. EMSA was performed with a 1- or 30-fold molar excess of unlabeled APO-Sp1/3 probe (competitor, lanes 6 and 7) or APO-Sp1/3mut probe (competitor mut., lanes 12 and 13). (B) Sp1- and Sp3-specific antibodies were added to the EMSA reactions resulting in a supershift (ss) of the respective antibody–protein–oligo complexes (lanes 3, 4, 7, 8, 11, 12). (C) ChIP assay was performed with DNA from A3.01 T cells. Immunoprecipitation was performed with antibodies against Sp1, Sp3 or actin. PCR primer pairs specific for the A3G promoter (upper panel) or the A3G gene (lower panel) were used. As positive controls, the sheared and cross-linked DNA before the immunoprecipitation step (input) or a plasmid carrying the target sequence (plasmid) was used as template."}

{"project":"events-check-again","denotations":[{"id":"T21335","span":{"begin":10,"end":13},"obj":"Protein"},{"id":"T21336","span":{"begin":18,"end":21},"obj":"Protein"},{"id":"T21337","span":{"begin":22,"end":26},"obj":"Binding"},{"id":"T21338","span":{"begin":22,"end":26},"obj":"Binding"},{"id":"T21339","span":{"begin":34,"end":40},"obj":"Entity"},{"id":"T21340","span":{"begin":56,"end":59},"obj":"Protein"},{"id":"T21341","span":{"begin":153,"end":156},"obj":"Protein"},{"id":"T21342","span":{"begin":279,"end":282},"obj":"Protein"},{"id":"T21343","span":{"begin":297,"end":300},"obj":"Protein"},{"id":"T21344","span":{"begin":301,"end":302},"obj":"Protein"},{"id":"T21345","span":{"begin":311,"end":314},"obj":"Protein"},{"id":"T21346","span":{"begin":315,"end":316},"obj":"Protein"},{"id":"T21347","span":{"begin":496,"end":499},"obj":"Protein"},{"id":"T21348","span":{"begin":500,"end":501},"obj":"Protein"},{"id":"T21349","span":{"begin":543,"end":546},"obj":"Protein"},{"id":"T21350","span":{"begin":547,"end":548},"obj":"Protein"},{"id":"T21351","span":{"begin":598,"end":601},"obj":"Protein"},{"id":"T21352","span":{"begin":607,"end":610},"obj":"Protein"},{"id":"T21353","span":{"begin":890,"end":893},"obj":"Protein"},{"id":"T21354","span":{"begin":895,"end":898},"obj":"Protein"},{"id":"T21355","span":{"begin":902,"end":907},"obj":"Protein"},{"id":"T21356","span":{"begin":943,"end":946},"obj":"Protein"},{"id":"T21357","span":{"begin":977,"end":980},"obj":"Protein"}],"relations":[{"id":"R17018","pred":"themeOf","subj":"T21335","obj":"T21337"},{"id":"R17019","pred":"themeOf","subj":"T21336","obj":"T21338"},{"id":"R17020","pred":"themeOf","subj":"T21339","obj":"T21337"},{"id":"R17021","pred":"partOf","subj":"T21339","obj":"T21340"},{"id":"R17022","pred":"themeOf","subj":"T21339","obj":"T21338"}],"text":"Figure 6. Sp1 and Sp3 bind to the GC-box present in the A3G promoter. (A) Nuclear extracts of A3.01 T cells were incubated with a 32P-labeled commercial Sp1 oligonucleotide probe (Sp1cons) or labeled probes homologous to the unmodified or mutated E2 region (see Figure 1) of the A3G promoter (APO-Sp1/3 and APO-Sp1/3mut). Protein–DNA complexes were separated by polyacrylamide electrophoresis and detected by autoradiography. EMSA was performed with a 1- or 30-fold molar excess of unlabeled APO-Sp1/3 probe (competitor, lanes 6 and 7) or APO-Sp1/3mut probe (competitor mut., lanes 12 and 13). (B) Sp1- and Sp3-specific antibodies were added to the EMSA reactions resulting in a supershift (ss) of the respective antibody–protein–oligo complexes (lanes 3, 4, 7, 8, 11, 12). (C) ChIP assay was performed with DNA from A3.01 T cells. Immunoprecipitation was performed with antibodies against Sp1, Sp3 or actin. PCR primer pairs specific for the A3G promoter (upper panel) or the A3G gene (lower panel) were used. As positive controls, the sheared and cross-linked DNA before the immunoprecipitation step (input) or a plasmid carrying the target sequence (plasmid) was used as template."}

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{"project":"bionlp-st-ge-2016-reference","denotations":[{"id":"T20690","span":{"begin":315,"end":316},"obj":"Protein"},{"id":"T20691","span":{"begin":496,"end":499},"obj":"Protein"},{"id":"T20692","span":{"begin":500,"end":501},"obj":"Protein"},{"id":"T20693","span":{"begin":543,"end":546},"obj":"Protein"},{"id":"T20694","span":{"begin":547,"end":548},"obj":"Protein"},{"id":"T20695","span":{"begin":598,"end":601},"obj":"Protein"},{"id":"T20696","span":{"begin":607,"end":610},"obj":"Protein"},{"id":"T20697","span":{"begin":890,"end":893},"obj":"Protein"},{"id":"T20698","span":{"begin":895,"end":898},"obj":"Protein"},{"id":"T20699","span":{"begin":902,"end":907},"obj":"Protein"},{"id":"T20700","span":{"begin":943,"end":946},"obj":"Protein"},{"id":"T20701","span":{"begin":977,"end":980},"obj":"Protein"},{"id":"T20679","span":{"begin":10,"end":13},"obj":"Protein"},{"id":"T20680","span":{"begin":18,"end":21},"obj":"Protein"},{"id":"T20681","span":{"begin":22,"end":26},"obj":"Binding"},{"id":"T20682","span":{"begin":22,"end":26},"obj":"Binding"},{"id":"T20683","span":{"begin":34,"end":40},"obj":"Entity"},{"id":"T20684","span":{"begin":56,"end":59},"obj":"Protein"},{"id":"T20685","span":{"begin":153,"end":156},"obj":"Protein"},{"id":"T20686","span":{"begin":279,"end":282},"obj":"Protein"},{"id":"T20687","span":{"begin":297,"end":300},"obj":"Protein"},{"id":"T20688","span":{"begin":301,"end":302},"obj":"Protein"},{"id":"T20689","span":{"begin":311,"end":314},"obj":"Protein"}],"relations":[{"id":"R16534","pred":"themeOf","subj":"T20679","obj":"T20681"},{"id":"R16535","pred":"themeOf","subj":"T20680","obj":"T20682"},{"id":"R16536","pred":"themeOf","subj":"T20683","obj":"T20681"},{"id":"R16537","pred":"partOf","subj":"T20683","obj":"T20684"},{"id":"R16538","pred":"themeOf","subj":"T20683","obj":"T20682"}],"namespaces":[{"prefix":"_base","uri":"http://bionlp.dbcls.jp/ontology/ge.owl#"}],"text":"Figure 6. Sp1 and Sp3 bind to the GC-box present in the A3G promoter. (A) Nuclear extracts of A3.01 T cells were incubated with a 32P-labeled commercial Sp1 oligonucleotide probe (Sp1cons) or labeled probes homologous to the unmodified or mutated E2 region (see Figure 1) of the A3G promoter (APO-Sp1/3 and APO-Sp1/3mut). Protein–DNA complexes were separated by polyacrylamide electrophoresis and detected by autoradiography. EMSA was performed with a 1- or 30-fold molar excess of unlabeled APO-Sp1/3 probe (competitor, lanes 6 and 7) or APO-Sp1/3mut probe (competitor mut., lanes 12 and 13). (B) Sp1- and Sp3-specific antibodies were added to the EMSA reactions resulting in a supershift (ss) of the respective antibody–protein–oligo complexes (lanes 3, 4, 7, 8, 11, 12). (C) ChIP assay was performed with DNA from A3.01 T cells. Immunoprecipitation was performed with antibodies against Sp1, Sp3 or actin. PCR primer pairs specific for the A3G promoter (upper panel) or the A3G gene (lower panel) were used. As positive controls, the sheared and cross-linked DNA before the immunoprecipitation step (input) or a plasmid carrying the target sequence (plasmid) was used as template."}

{"project":"bionlp-st-ge-2016-uniprot","denotations":[{"id":"T20936","span":{"begin":10,"end":13},"obj":"P08047"},{"id":"T20937","span":{"begin":18,"end":21},"obj":"Q02447"},{"id":"T20938","span":{"begin":56,"end":59},"obj":"Q9HC16"},{"id":"T20939","span":{"begin":153,"end":156},"obj":"P08047"},{"id":"T20940","span":{"begin":279,"end":282},"obj":"Q9HC16"},{"id":"T20941","span":{"begin":297,"end":300},"obj":"P08047"},{"id":"T20942","span":{"begin":311,"end":314},"obj":"P08047"},{"id":"T20943","span":{"begin":496,"end":499},"obj":"P08047"},{"id":"T20944","span":{"begin":543,"end":546},"obj":"P08047"},{"id":"T20945","span":{"begin":598,"end":601},"obj":"P08047"},{"id":"T20946","span":{"begin":607,"end":610},"obj":"Q02447"},{"id":"T20947","span":{"begin":890,"end":893},"obj":"P08047"},{"id":"T20948","span":{"begin":895,"end":898},"obj":"Q02447"},{"id":"T20949","span":{"begin":943,"end":946},"obj":"Q9HC16"},{"id":"T20950","span":{"begin":977,"end":980},"obj":"Q9HC16"}],"namespaces":[{"prefix":"_base","uri":"http://www.uniprot.org/uniprot/"}],"text":"Figure 6. Sp1 and Sp3 bind to the GC-box present in the A3G promoter. (A) Nuclear extracts of A3.01 T cells were incubated with a 32P-labeled commercial Sp1 oligonucleotide probe (Sp1cons) or labeled probes homologous to the unmodified or mutated E2 region (see Figure 1) of the A3G promoter (APO-Sp1/3 and APO-Sp1/3mut). Protein–DNA complexes were separated by polyacrylamide electrophoresis and detected by autoradiography. EMSA was performed with a 1- or 30-fold molar excess of unlabeled APO-Sp1/3 probe (competitor, lanes 6 and 7) or APO-Sp1/3mut probe (competitor mut., lanes 12 and 13). (B) Sp1- and Sp3-specific antibodies were added to the EMSA reactions resulting in a supershift (ss) of the respective antibody–protein–oligo complexes (lanes 3, 4, 7, 8, 11, 12). (C) ChIP assay was performed with DNA from A3.01 T cells. Immunoprecipitation was performed with antibodies against Sp1, Sp3 or actin. PCR primer pairs specific for the A3G promoter (upper panel) or the A3G gene (lower panel) were used. As positive controls, the sheared and cross-linked DNA before the immunoprecipitation step (input) or a plasmid carrying the target sequence (plasmid) was used as template."}