The GA-binding Protein Can Serve as Both an Activator and Repressor of ribosomal protein Gene Transcription

R Robert Genuario,Robert P Perry

doi:10.1074/jbc.271.8.4388

R Robert Genuario, Robert P Perry

Open Access

https://doi.org/10.1074/jbc.271.8.4388

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Abstract

The GA-binding protein (GABP), a heterodimeric transcription factor with widespread tissue distribution, has been found to be a strong positive regulator of several ribosomal protein (rp)-encoding genes. In such genes, e.g. the mouse rpL30 gene, the GABP-binding sites are located 40-80 base pairs upstream of the transcriptional start point. Potential GABP-binding sites are present in the promoters of numerous other rp genes, not only in upstream regions, but also in the immediate vicinity of the transcriptional start point. The mouse rpS16 gene is an example of the latter type. We demonstrate here that GABP binds to the rpS16 initiation region, and in so doing down-regulates rpS16 transcription both in vivo and in vitro. Supplementation of cell-free extracts with GABP inhibits transcription on rpS16 templates while concomitantly stimulating transcription on rpL30 templates. The repressive and stimulatory effects, which were proportional to the amount of GABP added, required both the GABP alpha subunit and either a beta1 or beta2 subunit. Mutations of the rpS16 GABP-binding sites that abolish binding increased rpS16 promoter activity in vivo and in vitro, whereas mutations that strengthen GABP binding caused a reduction in promoter activity. The binding of GABP to the rpS16 initiation region does not significantly affect the positioning of the transcriptional start points. Taken together with earlier studies, these new findings indicate that GABP can have a dual role as repressor or activator of rp gene transcription.

Highlights

GA-binding protein (GABP)1 ( called nuclear respiratory factor 2 (NRF-2) or E4 transcription factor 1 (E4TF1)) is a heteromeric transcription factor that has been shown to activate a number of genes, including those encoding the mouse ribosomal proteins L30 and L32 [1,2,3,4,5,6,7]
By constructing various mutated versions of these motifs and studying their effects on both GABP binding and rpS16 promoter function in vitro and in vivo, we further demonstrate that occupation of the binding site by either tetrameric or dimeric GABP causes a significant reduction in rpS16 promoter activity, apparently by interfering with the formation of the transcriptional initiation complex
The studies described above indicate that GABP can bind to the rpS16 initiation region, and in so doing modulate the activity of the rpS16 promoter

Summary

Introduction

GA-binding protein (GABP) ( called nuclear respiratory factor 2 (NRF-2) or E4 transcription factor 1 (E4TF1)) is a heteromeric transcription factor that has been shown to activate a number of genes, including those encoding the mouse ribosomal proteins L30 and L32 [1,2,3,4,5,6,7]. By constructing various mutated versions of these motifs and studying their effects on both GABP binding and rpS16 promoter function in vitro and in vivo, we further demonstrate that occupation of the binding site by either tetrameric or dimeric GABP causes a significant reduction in rpS16 promoter activity, apparently by interfering with the formation of the transcriptional initiation complex. Consistent with these observations, we found that supplementation of in vitro transcription reactions with GABP represses rpS16 transcription while concomitantly stimulating transcription of rpL30. This is the first report of a repressive role for GABP in transcriptional regulation

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