bhf-ucl The molecular basis of Rieger syndrome. Analysis of Pitx2 homeodomain protein activities. The molecular basis of Rieger syndrome. Analysis of Pitx2 homeodomain protein activities. J. Biol. Chem. (0021-9258) 1998 imex curation Amendt B.A., Sutherland L.B., Semina E.V., Russo A.F. Cardiac - Interactions involving cardiac related proteins Pitx2 has been entered as being of Human origin based on other work by the first author -see pmid 10490637. brad-amendt@uiowa.edu 05-Jan-2015: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro emsa electrophoretic mobility shift assay Gel retardation assay band shift band shift Gel retardation assay emsa autoradiography autoradiography Amendt B.A., Sutherland L.B., Semina E.V., Russo A.F. Cardiac - Interactions involving cardiac related proteins Pitx2 has been entered as being of Human origin based on other work by the first author -see pmid 10490637. brad-amendt@uiowa.edu Accepted 2014-NOV-10 by MGT J. Biol. Chem. (0021-9258) 1998 Only protein-protein interactions imex curation The molecular basis of Rieger syndrome. Analysis of Pitx2 homeodomain protein activities. The molecular basis of Rieger syndrome. Analysis of Pitx2 homeodomain protein activities. J. Biol. Chem. (0021-9258) 1998 imex curation Amendt B.A., Sutherland L.B., Semina E.V., Russo A.F. Cardiac - Interactions involving cardiac related proteins Pitx2 has been entered as being of Human origin based on other work by the first author -see pmid 10490637. brad-amendt@uiowa.edu 05-Jan-2015: Contacted by IntAct-Help. Only protein-protein interactions in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Amendt B.A., Sutherland L.B., Semina E.V., Russo A.F. Cardiac - Interactions involving cardiac related proteins Pitx2 has been entered as being of Human origin based on other work by the first author -see pmid 10490637. brad-amendt@uiowa.edu Accepted 2014-NOV-10 by MGT J. Biol. Chem. (0021-9258) 1998 Only protein-protein interactions imex curation pit-1 Pit-1 DNA binding element ss dna single stranded deoxyribonucleic acid ss DNA ss dna synthetic construct synthetic construct GATCCTGTCTTCCTGAATATGAATAAGAAATAAGGATC bcd_probe Drosophila bicoid site oligonucleotide ss dna single stranded deoxyribonucleic acid ss DNA ss dna synthetic construct synthetic construct GATCCGCACGGCCCATCTAATCCCGTGGGATC q99697-3 Pituitary homeobox 2 ARP1A PITX2 ARP1 RGS RIEG RIEG1 ALL1-responsive protein ARP1 Homeobox protein PITX2 Paired-like homeodomain transcription factor 2 RIEG bicoid-related homeobox transcription factor Solurshin protein protein human Homo sapiens Human METNCRKLVSACVQLEKDKSQQGKNEDVGAEDPSKKKRQRRQRTHFTSQQLQELEATFQRNRYPDMSTREEIAVWTNLTEARVRVWFKNRRAKWRKRERNQQAELCKNGFGPQFNGLMQPYDDMYPGYSYNNWAAKGLTSASLSTKSFPFFNSMNVNPLSSQSMFSPPNSISSMSMSSSMVPSAVTGVPGSSLNSLNNLNNLSSPSLNSAVPTPACPYAPPTPPYVYRDTCNSSLASLRLKAKQHSSFGYASVQNPASNLSACQYAVDRPV Variant in position: 58:F->L (in ASGD4). 45D2ADA6DF03FF65 prd Prd ss dna single stranded deoxyribonucleic acid ss DNA ss dna synthetic construct synthetic construct GATCCGCACGGCCCATCTTTGACGTCGTGGGATC pit1_rat Pituitary-specific positive transcription factor 1 Pou1f1 Ghf-1 Pit-1 Pit1 Growth hormone factor 1 protein protein rat Rattus norvegicus (Rat) Rat MSCQPFTSADTFIPLNSDASAALPLRMHHSAAEGLPASNHATNVMSTATGLHYSVPSCHYGNQPSTYGVMAGTLTPCLYKFPDHTLSHGFPPLHQPLLAEDPTASEFKQELRRKSKLVEEPIDMDSPEIRELEQFANEFKVRRIKLGYTQTNVGEALAAVHGSEFSQTTICRFENLQLSFKNACKLKAILSKWLEEAEQVGALYNEKVGANERKRKRRTTISIAAKDALERHFGEHSKPSSQEIMRMAEELNLEKEVVRVWFCNRRQREKRVKTSLNQSLFSISKEHLECR 2651876B2F7668A1 nkx-2.1 Nkx-2.1 ss dna single stranded deoxyribonucleic acid ss DNA ss dna synthetic construct synthetic construct GATCCCACTGCCCAGTCAAGTGTTCGGATC ftz Ftz ss dna single stranded deoxyribonucleic acid ss DNA ss dna synthetic construct synthetic construct GATCCGCACGGCCCAGTTAATGGCGTGGGATC mu 9 Mu 9 ss dna single stranded deoxyribonucleic acid ss DNA ss dna synthetic construct synthetic construct GATCCGCACGGCCCAGTTAATATCGTGGGATC bcd-pitx2-2 1 5 unspecified role unspecified role prey prey ecoli Escherichia coli (strain K12) 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct direct interaction direct interaction Fig. 1C, 2A, 2C, 6 Pit-1 (P10037) bcd-pitx2-3 1 5 unspecified role unspecified role prey prey ecoli Escherichia coli (strain K12) 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct 3 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct physical association physical association Fig. 2A, 2B Unlabeled Pit-1 DNA binding element caused 34% inhibition. bcd-pitx2-5 1 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 15:40:52 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 19:46:52 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 10 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 3A, 3B The mutant Pitx2 T68P protein showed an overall increase in binding to all of the competitor DNAs (except the unlabeled bicoid DNA). The ability of these nonbicoid DNA competitors to bind Pitx2 T68P demonstrates a slight loss of binding specificity by this mutant. pitx2-pou1f1-2 2 5 unspecified role unspecified role bait bait n-terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range ecoli Escherichia coli (strain K12) 7 unspecified role unspecified role prey prey direct interaction direct interaction Fig. 7 O93385-2 and P10037 increase expression at RNA level of ENSRNOG00000017374. bcd-pitx2-6 1 5 unspecified role unspecified role prey prey ecoli Escherichia coli (strain K12) 4 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 2A, 2B Unlabeled bicoid DNA (Bcd) caused more than 85% inhibition. bcd-pitx2-7 1 5 unspecified role unspecified role prey prey ecoli Escherichia coli (strain K12) 6 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 2A, 2B Unlabeled Prd caused less than 20% inhibition. bcd-pitx2-8 1 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 18:33:32 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 18:07:56 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 4 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 3A, 3B The mutant Pitx2 T68P protein showed an overall increase in binding to all of the competitor DNAs (except the unlabeled bicoid DNA). The ability of these nonbicoid DNA competitors to bind Pitx2 T68P demonstrates a slight loss of binding specificity by this mutant. bcd-pitx2-9 1 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 14:37:18 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 19:52:58 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 6 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 3A, 3B The mutant Pitx2 T68P protein showed an overall increase in binding to all of the competitor DNAs (except the unlabeled bicoid DNA). The ability of these nonbicoid DNA competitors to bind Pitx2 T68P demonstrates a slight loss of binding specificity by this mutant. bcd-pitx2-10 1 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 15:37:09 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 16:26:02 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct 3 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct physical association physical association Fig. 3A, 3B The mutant Pitx2 T68P protein showed an overall increase in binding to all of the competitor DNAs (except the unlabeled bicoid DNA). The ability of these nonbicoid DNA competitors to bind Pitx2 T68P demonstrates a slight loss of binding specificity by this mutant. bcd-pitx2-11 1 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 17:45:45 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 16:32:40 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 9 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 3A, 3B The mutant Pitx2 T68P protein showed an overall increase in binding to all of the competitor DNAs (except the unlabeled bicoid DNA). The ability of these nonbicoid DNA competitors to bind Pitx2 T68P demonstrates a slight loss of binding specificity by this mutant. bcd-pitx2-13 1 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 20:21:57 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 16:05:14 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct 8 competitor competitor competitor unspecified role unspecified role synthetic construct synthetic construct physical association physical association Fig. 3A, 3B The mutant Pitx2 T68P protein showed an overall increase in binding to all of the competitor DNAs (except the unlabeled bicoid DNA). The ability of these nonbicoid DNA competitors to bind Pitx2 T68P demonstrates a slight loss of binding specificity by this mutant. bcd-pitx2-14 1 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 17:09:50 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 19:45:35 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct direct interaction direct interaction Fig. 1C, 3A, 3C, 6 Pit-1 (P10037) EMSAs demonstrated binding of Pitx2 and Pitx2 T68P mutant to the bicoid element (Fig. 1C). Binding to Pitx2 L54Q mutant was not detected most likely because of protein instability. bcd-pitx2-17 1 5 unspecified role unspecified role prey prey ecoli Escherichia coli (strain K12) 7 stimulator stimulator unspecified role unspecified role 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 6 Pit-1 increased the amount of Pitx2-DNA bound complex approximately 2–3-fold. The Pit-1-enhanced binding of Pitx2 was competed by unlabeled bicoid DNA bcd-pitx2-18 1 7 stimulator stimulator unspecified role unspecified role 5 unspecified role unspecified role prey prey Q99697-3:p.Thr68Pro mutation mutation certain certain sequence position certain certain certain sequence position certain T P Wed May 02 16:58:12 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr68pro' Tue Jun 30 17:53:56 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr68Pro' ecoli Escherichia coli (strain K12) 4 unspecified role unspecified role bait bait region 32p radiolabel 32p radiolabel P32 32P 32P P32 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined synthetic construct synthetic construct physical association physical association Fig. 6 Pit-1 had a 2-3 fold stimulatory effect on the binding of the mutant Pitx2 T68P to the bicoid probe.