TY - JOUR
T1 - Isolation and characterization of the binding sequences for the product of the arabidopsis floral homeotic gene AGAMOUS
AU - Huang, Hai
AU - Mizukami, Yukiko
AU - Hu, Yi
AU - Ma, Hong
N1 - Funding Information:
We would like to thank E.Grotewold for helpful advice and discussion on DNA-binding experiments, C.Flanagan for the antiserum used in the western analysis of AG, D.Grueneberg for the gift of SRF and advice, S.Bell for advice on footprinting analysis, M.Tudor for help with initial experiments, and E.Grotewold, D.Grueneberg, C.Flanagan, P.Rubinelli, C.Weiss and the anonymous reviewers for helpful comments on this manuscript. This work was supported by funds from the Cold Spring Harbor Laboratory Robertson Fund and by a grant from the National Science Foundation (DCB-9105260) to H.M.
PY - 1993/10/11
Y1 - 1993/10/11
N2 - The Arabidopsis floral homeotic geneAGAMOUS (AG) is required for normal flower development. The deduced AG protein contains a region which shares substantial sequence similarity with the DNA-binding domains of known transcription factors, SRF (human) and MCM1 (yeast). Therefore, it is likely that AG is also a DNA-binding protein regulating transcription of floral genes. We describe here several experiments to characterize AG-DNA binding in vitro. We show that AG indeed binds a DNA sequence matching the consensus of SRF targets. Further, we have selected the AGbinding sequences from a pool of random oligonucleotides, and deduced an AG-binding consensus sequence of TT(A/T)CC(A/T)(A/t)2(T/A)NNGG(-G)(A/t)2. We have demonstrated that AG binds to the consensus region of three of the oligonucleotides by footprinting analysis. Finally, we have examined AG′s relative binding affinity for different sequences, as compared to SRF, by gel mobility shift analysis. Our results indicate that AG is a sequence-specific DNA-binding protein, and that the AG-binding consensus sequence is similar to those of MCM1 and SRF.
AB - The Arabidopsis floral homeotic geneAGAMOUS (AG) is required for normal flower development. The deduced AG protein contains a region which shares substantial sequence similarity with the DNA-binding domains of known transcription factors, SRF (human) and MCM1 (yeast). Therefore, it is likely that AG is also a DNA-binding protein regulating transcription of floral genes. We describe here several experiments to characterize AG-DNA binding in vitro. We show that AG indeed binds a DNA sequence matching the consensus of SRF targets. Further, we have selected the AGbinding sequences from a pool of random oligonucleotides, and deduced an AG-binding consensus sequence of TT(A/T)CC(A/T)(A/t)2(T/A)NNGG(-G)(A/t)2. We have demonstrated that AG binds to the consensus region of three of the oligonucleotides by footprinting analysis. Finally, we have examined AG′s relative binding affinity for different sequences, as compared to SRF, by gel mobility shift analysis. Our results indicate that AG is a sequence-specific DNA-binding protein, and that the AG-binding consensus sequence is similar to those of MCM1 and SRF.
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U2 - 10.1093/nar/21.20.4769
DO - 10.1093/nar/21.20.4769
M3 - Article
C2 - 7901838
AN - SCOPUS:0027524204
SN - 0305-1048
VL - 21
SP - 4769
EP - 4776
JO - Nucleic acids research
JF - Nucleic acids research
IS - 20
ER -