RT Journal Article
SR Electronic
T1 RNA- and DNA-binding proteins generally exhibit direct transfer of polynucleotides: Implications for target site search
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2022.11.30.518605
DO 10.1101/2022.11.30.518605
A1 Wayne O. Hemphill
A1 Calvin K. Voong
A1 Regan Fenske
A1 James A. Goodrich
A1 Thomas R. Cech
YR 2022
UL http://biorxiv.org/content/early/2022/11/30/2022.11.30.518605.abstract
AB We previously demonstrated that the PRC2 chromatin-modifying enzyme exhibits the ability to directly transfer between RNA and DNA without a free-enzyme intermediate state. Simulations suggested that such a direct transfer mechanism may be generally necessary for RNA to recruit proteins to chromatin, but the prevalence of direct transfer capability is unknown. Herein, we used fluorescence polarization assays and observed direct transfer for several well-characterized nucleic acid-binding proteins: three-prime repair exonuclease 1 (TREX1), heterogeneous nuclear ribonucleoprotein U, Fem-3-binding factor 2, and MS2 bacteriophage coat protein. For TREX1, the direct transfer mechanism was additionally interrogated by single molecule assays, and the data suggest that direct transfer occurs through an unstable ternary intermediate with partially associated ligands. Generally, direct transfer could allow many DNA- and RNA-binding proteins to conduct a one-dimensional search for their target sites. Furthermore, presumably long-lived protein-polynucleotide complexes might instead be readily replaced by other protein-polynucleotide complexes in vivo.Significance Classically, the lifetime of a protein-ligand complex is presumed to be an intrinsic property, unaffected by competitor molecules in free solution. By contrast, a few oligomeric nucleic acid binding proteins have been observed to exchange competing ligands in their binding sites, and consequently their lifetimes decrease with competitor concentration. Our findings indicate that this “direct transfer” is a more general property of nucleic acid binding proteins. This suggests that many DNA- and RNA-binding proteins can reduce the dimensionality of their search for their target sites by intramolecular direct transfer to nucleosome DNA, instead of relying entirely on three-dimensional diffusion, and it suggests that their mean complex lifetimes in vivo can be regulated by the concentration of free ligand molecules.Competing Interest StatementT.R.C declares consulting status for Storm Therapeutics, Eikon Therapeutics, and SomaLogic. W.O.H. declares the filing of U.S. Provisional Application No. 62/706,167 Trex1 Inhibitors and Uses Thereof. The authors have no other competing interests to declare.