suGF1 binds in the major groove of its oligo(dG).oligo(dC) recognition sequence and is excluded by a positioned nucleosome core.

Abstract

We have elsewhere reported the purification of a poly(dG) poly(dC)-binding nuclear protein (suGFl) from sea urchin embryos (J. Hapgood and D. Patterton, Mol. Cell. Biol. 14:this issue, 1994). We proposed that suGFl may be a member of a family of G-string factors involved in developmental gene regulation, possibly via alterations in chromatin structure. In this article, we characterize the binding of purified suGFl to 11 contiguous Gs in the H1-H4 intergenic region of a sea urchin early histone gene battery in vitro. It is shown that suGFl-DNA binding is dependent on ionic strength and requires divalent cations. Purified suGFl forms discrete protein-DNA multimers, consistent with suGFl-suGFl interactions. In a model for the suGFl-DNA complex derived from our footprinting and methylation interference data, suGFl contacts the Gs in the major groove as well as one of the bordering phosphate backbones. The data are consistent with the direction of curvature of the DNA in the suGFl-DNA complex being the same as that preferred by the free DNA and exhibited by the DNA when bent around a positioned nucleosome core in vitro. However, on the basis of steric considerations, the binding of suGFl and that of the histone octamer are predicted to be mutually exclusive.We show that suGFl is indeed unable to bind to the G string when occupied by a histone octamer located in the major in vitro positioning frame in the H1-H4 intergenic region.

Purification of an oligo(dG).oligo(dC)-binding sea urchin nuclear protein, suGF1: a family of G-string factors involved in gene regulation during development.

“…It is not possible with the available data to deduce whether these results reflect a difference between the DNAbinding specificities of BGP1 and suGF1. BGP1 was shown to require the presence of Zn2+ for DNA binding (22), whereas we were unable to show a Zn2+ requirement for suGF1 (25).…”

“…A single DNase I footprint was also detected on the Crick strand at the same position (data not shown). We present elsewhere a detailed analysis of the purified suGF1-DNA interaction in the H1-H4 intergenic region by methylation interference and footprinting with chemical and enzymatic probes (25).…”

Identification of a key regulatory element for the basal activity of the human insulin-like growth factor II gene promoter P3

“…Thirdly, proteins 4 and 5 are unable to bind to single-stranded DNA (results not shown). This ds-DNA-binding specificity was also shown to exist for the G-string-binding factor suGF1 [26].…”

“…Interestingly, the requirement for bivalent ions has also been shown for the G-string-binding factor BGP1, which needs Zn# + ions for its DNA-binding activity [23]. Furthermore it was shown that the sea-urchin G-string-binding factor suGF1 is also tightly complexed with a cation [26]. Thirdly, proteins 4 and 5 are unable to bind to single-stranded DNA (results not shown).…”

“…In the chicken, two G-string-element-containing promoters have been described that are bound by the chicken erythrocyte protein called β-globin protein 1, BGP 1 [23,24]. In the sea urchin (Psammechinus miliaris) the purification of another G-stringbinding protein of 59.5 kDa, called sea-urchin G-string factor 1 (suGF1), has been described [25,26]. The promoter of the celllineage-specific gene LpS-β of the sea urchin Lytechinus pictus also contains a G-string element dG ' dC that can be bound by suGF1 [27].…”