Repeat units from a maize rDNA external spacer region exhibit DNA curvature and interact with high-mobility-group proteins

Abstract

The 3-kb external spacer from a maize (Zea mays L. cv. A619) nuclear rRNA gene unit which contains nine highly homologous 200-bp repeat elements was found to include a region with DNA-curvature properties. The centre of curvature was localized within repeats 5 and 6 using a circular permutation assay. A 60-bp-long subfragment of this region was found to interact with nuclear proteins, including high-mobility-group (HMG) proteins, and with the maize HMGa protein synthesized in Escherichia coli from a recombinant plasmid. The potential influence of the binding of the HMG proteins on the conformation of this subfragment was studied with a permutation assay based on a bending vector.

An intrinsically bent region upstream of the transcription start site of the rRNA genes of Arabidopsis thaliana interacts with an HMG-related protein

“…By use of a circular permutation assay this bending centre was localized to position -284 to -256 upstream of the transcription start site. Intrinsic D N A bending was also found in the external spacer region (ESR) of maize r D N A [ 9] and in the NTS of pea r D N A [ 11 ]. As first proposed by Koo et al [14], the sequence requirements for D N A bending are dA°dT tracts.…”

“…It is highly probable that such transcription factors exist also in the plant kingdom. Indeed, various proteins were found to bind specific sequences upstream of the TSS of different plant rRNA genes [9,12,20,25,33]. However, n o n e o f these proteins has been purified so far.…”

“…Griess et al [9] reported a bent DNA structure upstream of the TS S of the rRNA genes of maize, that is bound by an H M G protein. The authors speculate, that bent DNA conformations stabilize the binding of H M G proteins by contracting the minor groove, the site of D N A -H M G protein interaction.…”

Ribosomal transcription units integrated via T‐DNA transformation associate with the nucleolus and do not require upstream repeat sequences for activity in Arabidopsis thaliana

“…The system should also complement a number of efforts to define parts of the intergenic region important for rRNA synthesis which have so far relied on gel mobility shift assays to detect DNA-protein interactions (Ashapkin et al, 1995;Echeverria et aL, 1992;Echeverria et aL, 1994;Griess et aL, 1993;Kneidl et aL, 1995;Nakajima et al, 1992;Zentgraf and Hemleben, 1992). Previously, comparative analysis of rDNA intergenic regions has lead to models (Cordesse etal., 1993;Gr0ndler etal., 1991;Hemleben and Zentgraf, 1994;King et aL, 1993) which can now be tested for the model plant species A. thaliana.…”

Comparative analysis of chromosomal HMG proteins from monocotyledons and dicotyledons

“…Wheat and barley HMG proteins have been reported to be preferentially associated with actively transcribed chromatin [18,25]. Some HMG proteins from maize, soybean and wheat have been shown by in vitro studies to bind to A/T-rich stretches in the 5'-flanking region of polymerase I and II genes [ 10,12,17,20]. In maize, preferential binding of the HMGa and HMGb proteins to the CCAAT and TATA boxes of the P2 promoter of the zein storage protein gene pMS1 was observed [8].…”