Plant Chromosomal HMGB Proteins Efficiently Promote the Bacterial Site-Specific β-Mediated Recombination in Vitro and in Vivo

Abstract

In the presence of an accessory DNA bending protein, the bacterial site-specific beta recombinase catalyzes resolution and DNA inversion. Five different maize high mobility group B (HMGB) proteins were examined for their potential to facilitate beta recombination in vitro using DNA substrates with different intervening distances (73-913 bp) between two directly oriented recombination (six) sites. All analyzed HMGB proteins (HMGB1 to HMGB5) could promote beta recombination, but depending on the DNA substrate with different efficiencies. The HMGB1 protein displayed an activity comparable to that of the natural promoting protein Hbsu, whereas the other HMGB proteins were less effective. Phosphorylation of the HMGB1 protein resulted in an increased efficiency of HMGB1 to promote beta recombination. Analyses of DNA substrates with closely spaced six sites demonstrated that in the presence of HMGB1 the recombination rate was correlated to the distance between the six sites, but independent of the helical orientation of the six sites. Using a Bacillus subtilis strain defective in Hbsu, the coexpression of beta recombinase and HMGB1 (or a truncated HMGB1 derivative) revealed that a plant HMG-box domain protein is sufficient for assisting beta to catalyze recombination in vivo. Our results using beta recombination as a model system suggest that the various plant HMGB proteins (and their posttranslationally modified versions) have the potential of forming a repertoire of different DNA structures, which is compatible with the idea that the HMGB proteins can act as architectural factors in a variety of nucleoprotein structures.

The HMG-box: a versatile protein domain occurring in a wide variety of DNA-binding proteins

“…The efficiency of various maize HMGB-type proteins in promoting the formation of a nucleoprotein complex was tested using a site-specific recombination reaction as a model. In this assay, the HMGB-type proteins (depending on the type of DNA substrate) stimulated the b recombination reaction to different extents, indicating that the proteins differ in their ability to assist the assembly of nucleoprotein structures [113]. Similarly, the wheat and maize HMGBtype proteins have different potential to facilitate the binding of certain transcription factors to their target DNA sites [114,115].…”

“…Plant proteins containing HMG-box domain(s).Structure-specific DNA binding, bending[102] Structure-specific DNA binding, bending, b recomb., NL[98,101,103,113] Structure-specific DNA binding, bending, b recomb., NL[98,101,103,113] Structure-specific DNA binding, bending, b recomb., NL[98,101,103,113] Structure-specific DNA binding, bending, b recomb., NL[98,101,103,113] Stimulation of transcription factor Dof2, b recomb [113,114,116]. Structure-specific DNA binding, bending, NL[122,124] a Published name of protein or * name used in Plant Chromatin Database (http://www.chromdb.org/) with sequences from Arabidopsis thaliana (At), Zea mays (Zm), Oryza sativa (Os) and Populus trichocarpa (Pt).…”

Plant Proteins Containing High Mobility Group Box DNA-Binding Domains Modulate Different Nuclear Processes

“…The potential of plant HMGB proteins to promote the formation of specific nucleoprotein structures is evident from their stimulatory architectural role in site-specific recombination reactions, which was observed in vitro and in vivo (Stemmer et al, 2002). Moreover, HMGB proteins can facilitate the binding of certain plant transcription factors (basic Leu zipper, DNA binding with one finger [DOF]) to their DNA recognition sites by direct interaction (in several instances via their HMG-boxes) with these factors (Schultz et al, 1996;Yanagisawa, 1997).…”

Functionality of the β/six Site-Specific Recombination System in Tobacco and Arabidopsis: A Novel Tool for Genetic Engineering of Plant Genomes

“…In Bacillus subtilis, b-mediated recombination is strictly dependent on the presence of the host factor Hbsu (an HU-like protein) that facilitates the correct assembly of the synaptic complex [33,34]. A plastid-encoded HU-like protein from the cryptomonad alga Guillardia theta [35], as well as chromatin-associated HMGB proteins from mammals [36] and plants [37,38] can efficiently substitute for the Hbsu protein assisting b recombination. All the plant HMGB proteins tested could facilitate in vitro b recombination, albeit (depending on the DNA substrate) with different efficiency.…”