DNA Binding Specificity Studies of Four ETS Proteins Support an Indirect Read-out Mechanism of Protein-DNA Recognition* 210

Members of the ETS family of transcription factors are involved in several developmental and physiological processes, and, when overexpressed or misexpressed, can contribute to a variety of cancers. Each family member has a conserved DNA-binding domain that recognizes DNA sequences containing a G-G-A trinucleotide. Discrimination between potential ETS-binding sites appears to be governed by both the nucleotides flanking the G-G-A sequence and protein-protein interactions. We have used an adaptation of the “length-encoded multiplex” approach (Desjarlais, J. R., and Berg, J. M. (1994) Proc. Natl. Acad. Sci. U. S. A.91, 11099–11103) to define DNA binding specificities for four ETS proteins: Fli-1, SAP-1, PU.1, and TEL. Our results support a model in which cooperative effects among neighboring bases flanking the central G-G-A site contribute to the formation of stable ETS/DNA complexes. These results are consistent with a mechanism for specific DNA binding that is partially governed by an indirect read-out of the DNA sequence, in which a sequence-specific DNA conformation is sensed or induced.

[1]  B. Wasylyk,et al.  The Ets family of transcription factors. , 1993, European journal of biochemistry.

[2]  N Muthusamy,et al.  The Ets-1 transcription factor is required for the development of natural killer cells in mice. , 1998, Immunity.

[3]  A. Sharrocks,et al.  The ETS-domain transcription factor family. , 1997, Nature reviews. Molecular cell biology.

[4]  E. Scott,et al.  Requirement of transcription factor PU.1 in the development of multiple hematopoietic lineages. , 1994, Science.

[5]  R. Dickerson,et al.  DNA structure from A to Z. , 1992, Methods in enzymology.

[6]  Y Fujiwara,et al.  The TEL/ETV6 gene is required specifically for hematopoiesis in the bone marrow. , 1998, Genes & development.

[7]  N Assa-Munt,et al.  Mutants of ETS domain PU.1 and GGAA/T recognition: Free energies and kinetics , 1999, Protein science : a publication of the Protein Society.

[8]  F. Moreau-Gachelin,et al.  DNA binding specificities of Spi-1/PU.1 and Spi-B transcription factors and identification of a Spi-1/Spi-B binding site in the c-fes/c-fps promoter. , 1995, Oncogene.

[9]  A. Gronenborn,et al.  Correction of the NMR structure of the ETS1/DNA complex , 1997, Journal of biomolecular NMR.

[10]  S. McKnight,et al.  Specificities of protein-protein and protein-DNA interaction of GABP alpha and two newly defined ets-related proteins. , 1992, Genes & development.

[11]  O. Bernard,et al.  TEL Is a Sequence-specific Transcriptional Repressor* , 1999, The Journal of Biological Chemistry.

[12]  J. Green,et al.  Differential expression of ets-1 and ets-2 proto-oncogenes during murine embryogenesis. , 1994, Oncogene.

[13]  C. Thummel,et al.  Molecular interactions within the ecdysone regulatory hierarchy: DNA binding properties of the Drosophila ecdysone-inducible E74A protein , 1990, Cell.

[14]  B. Graves,et al.  Interaction of murine ets-1 with GGA-binding sites establishes the ETS domain as a new DNA-binding motif. , 1992, Genes & development.

[15]  J. Ghysdael,et al.  Identification of nucleotide preferences in DNA sequences recognised specifically by c-Ets-1 protein. , 1992, Nucleic acids research.

[16]  L. McIntosh,et al.  Solution structure of the ETS domain from murine Ets‐1: a winged helix‐turn‐helix DNA binding motif. , 1996, The EMBO journal.

[17]  A. Mårtensson,et al.  Spi-C, a Novel Ets Protein That Is Temporally Regulated during B Lymphocyte Development* , 1999, The Journal of Biological Chemistry.

[18]  M. Abe,et al.  ETS‐1 converts endothelial cells to the angiogenic phenotype by inducing the expression of matrix metalloproteinases and integrin β3 , 1999, Journal of cellular physiology.

[19]  A. Amores,et al.  Molecular characterization of the zebrafish PEA3 ETS-domain transcription factor , 1998, Oncogene.

[20]  R. Kodandapani,et al.  A new pattern for helix–turn–helix recognition revealed by the PU.l ETS–domain–DNA complex , 1996, Nature.

[21]  A. J. Valente,et al.  Critical Flanking Sequences of PU.1 Binding Sites in Myeloid-specific Promoters* , 1999, The Journal of Biological Chemistry.

[22]  Cynthia Wolberger,et al.  The Structure of GABPα/β: An ETS Domain- Ankyrin Repeat Heterodimer Bound to DNA , 1998 .

[23]  M. Abe,et al.  Ets‐1 regulates angiogenesis by inducing the expression of urokinase‐type plasminogen activator and matrix metalloproteinase‐1 and the migration of vascular endothelial cells , 1996, Journal of cellular physiology.

[24]  Hydroxyl radical footprinting of DNA complexes of the ets domain of PU.1 and its comparison to the crystal structure. , 1998, Biochemistry.

[25]  Ronen Marmorstein,et al.  Structure of the Elk-1–DNA complex reveals how DNA-distal residues affect ETS domain recognition of DNA , 2000, Nature Structural Biology.

[26]  J R Desjarlais,et al.  Length-encoded multiplex binding site determination: application to zinc finger proteins. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[27]  R. Fisher,et al.  High-affinity DNA-protein interactions of the cellular ETS1 protein: the determination of the ETS binding motif. , 1991, Oncogene.

[28]  M. Tymms,et al.  The Ets1 transcription factor is widely expressed during murine embryo development and is associated with mesodermal cells involved in morphogenetic processes such as organ formation. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[29]  W. Leonard,et al.  Importance of low affinity Elf-1 sites in the regulation of lymphoid- specific inducible gene expression , 1996, The Journal of experimental medicine.

[30]  R. Berger,et al.  The TEL gene products: nuclear phosphoproteins with DNA binding properties , 1997, Oncogene.

[31]  J. Taylor,et al.  A role for the ETS domain transcription factor PEA3 in myogenic differentiation , 1997, Molecular and cellular biology.

[32]  F. Studier.  Use of bacteriophage T7 lysozyme to improve an inducible T7 expression system. , 1991, Journal of molecular biology.

[33]  A. Sharrocks,et al.  Molecular characterization of a zebrafish TCF ETS-domain transcription factor , 1999, Oncogene.

[34]  L. J. Maher,et al.  Role of Asymmetric Phosphate Neutralization in DNA Bending by PU.1* , 1997, The Journal of Biological Chemistry.

[35]  R. Meadows,et al.  Solution structure of the ets domain of Fli-1 when bound to DNA , 1994, Nature Structural Biology.

[36]  A. Sharrocks,et al.  The ETS-domain transcription factors Elk-1 and SAP-1 exhibit differential DNA binding specificities. , 1995, Nucleic acids research.

[37]  C. Thompson,et al.  The FLI-1 and chimeric EWS-FLI-1 oncoproteins display similar DNA binding specificities. , 1994, The Journal of biological chemistry.

[38]  R. Marmorstein,et al.  Structures of SAP-1 bound to DNA targets from the E74 and c-fos promoters: insights into DNA sequence discrimination by Ets proteins. , 1998, Molecular cell.