Intron 1 Elements Promote Erythroid-specific GATA-1 Gene Expression

• Published in: The Journal of biological chemistry 2000-07, Vol.275 (30), p.22969-22977
• Main Authors: Seshasayee, Dhaya, Geiger, Justin N., Gaines, Peter, Wojchowski, Don M.
• Format: Article
• Language: English
• Subjects: Base Sequence; Cell Line; DNA; DNA Primers; DNA-Binding Proteins - genetics; Erythrocytes - metabolism; Erythroid-Specific DNA-Binding Factors; GATA-1 protein; GATA1 Transcription Factor; Gene Expression Regulation - genetics; Introns; Molecular Sequence Data; Transcription Factors - genetics; Transcription, Genetic - genetics
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M002931200

The zinc finger protein GATA-1 functions in a concentration-dependent fashion to activate the transcription of erythroid and megakaryocytic genes. Less is understood, however, regarding factors that regulate the GATA-1 gene. Presently elements within intron 1 are shown to markedly affect its erythroid-restricted transcription. Within a full-length 6.8-kilobaseGATA-1 gene construct (G6.8-Luc) the deletion of a central subdomain of intron 1 inhibited transcription ≥10-fold in transiently transfected erythroid SKT6 cells, and likewise inhibited high-level transcription in erythroid FDCW2ER-GATA1 cells. In parental myeloid FDCER cells, however, low-level transcription was largely unaffected by intron 1 deletions. Within intron 1, repeated GATA and Ap1 consensus elements in a central region are described which when linked directly to reporter cassettes promote transcription in erythroid SKT6 and FDCER-GATA1 cells at high rates. Moreover, GATA-1 activated transcription from this subdomain in 293 cells, and in SKT6 cells this subdomain footprinted in vivo. For stably integrated GFP reporter constructs in erythroid SKT6 cells, corroborating results were obtained. Deletion of intronic GATA and Ap1 motifs abrogated the activity of G6.8-pEGFP; activity was decreased by 43 and 56%, respectively, by the deletion of either motif; and the above 1800-base pair region of intron 1 per se was transcribed at rates uniformly greater than G6.8-pEGFP. Also described is the differential utilization of exons 1a and 1b among primary erythromegakaryocytic and myeloid cells.