Biology subjects

Salthouse, D.

Publications and source records attributed to Salthouse, D..

3 recordsLinked to original sources

bioRxiv2026

A chemoinformatics-guided platform for efficient discovery of RNA-binding small molecules: Proof-of-concept for myotonic dystrophy type 1

Structured RNAs cause human diseases but remain challenging to target selectively with small molecules. Here, we report a chemoinformatics-guided discovery framework that integrates fingerprint-based molecular design, experimental validation, and mechanistic profiling to identify small molecules that bind highly structured, disease-associated RNAs. Using an RNA-binder fingerprint derived from known ligands, a Tversky similarity screen of >8 million compounds yielded a 150-member library enriched in chemical space for RNA-active scaffolds. Target engagement and cell-based assays identified multiple selective ligands for the pathogenic expanded triplet repeat, r(CUG)exp, that causes myotonic dystrophy type 1 (DM1) by binding and sequestering the RNA-binding protein muscleblind-like 1 (MBNL1). Biophysical and single-molecule analyses revealed that the small molecules bind the 1x1 nucleotide U/U internal loops formed when r(CUG)exp folds, partially block MBNL1 binding, and modulate RNA folding equilibria. Two optimized scaffolds rescued MBNL1-dependent splicing in patient-derived myotubes with micromolar potency and minimal cytotoxicity. This study establishes a generalizable, data-driven platform for discovering drug-like RNA-binding lead small molecules and demonstrates its application to the toxic repeat expansion RNA underlying DM1. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=97 SRC="FIGDIR/small/723748v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@1a87b41org.highwire.dtl.DTLVardef@340a14org.highwire.dtl.DTLVardef@81b583org.highwire.dtl.DTLVardef@1b3ba14_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

bioinformatics↗

bioRxiv2024

NMR structures and magnetic force spectroscopy studies of small molecules binding to models of an RNA CAG repeat expansion

RNA repeat expansions fold into stable structures and cause microsatellite diseases such as Huntingtons disease (HD), myotonic dystrophy type 1 (DM1), and spinocerebellar ataxias (SCAs). The trinucleotide expansion of r(CAG), or r(CAG)exp, causes both HD and SCA3, and the RNAs toxicity has been traced to its translation into polyglutamine (polyQ; HD) as well as aberrant pre-mRNA alternative splicing (SCA3 and HD). Previously, a small molecule, 1, was discovered that binds to r(CAG)exp and rescues aberrant pre-mRNA splicing in patient-derived fibroblasts by freeing proteins bound to the repeats. Here, we report the structures of single r(CAG) repeat motif (5CAG/3GAC where the underlined adenosines form a 1x1 nucleotide internal loop) in complex with 1 and two other small molecules via nuclear magnetic resonance (NMR) spectroscopy combined with simulated annealing. Compound 2 was designed based on the structure of 1 bound to the RNA while 3 was selected as a diverse chemical scaffold. The three complexes, although adopting different 3D binding pockets upon ligand binding, are stabilized by a combination of stacking interactions with the internal loops closing GC base pairs, hydrogen bonds, and van der Waals interactions. Molecular dynamics (MD) simulations performed with NMR-derived restraints show that the RNA is stretched and bent upon ligand binding with significant changes in propeller-twist and opening. Compound 3 has a distinct mode of binding by insertion into the helix, displacing one of the loop nucleotides into the major groove and affording a rod-like shape binding pocket. In contrast, 1 and 2 are groove binders. A series of single molecule magnetic force spectroscopy studies provide a mechanistic explanation for how bioactive compounds might rescue disease-associated cellular phenotypes. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/608150v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@145398org.highwire.dtl.DTLVardef@7c37caorg.highwire.dtl.DTLVardef@132cbbeorg.highwire.dtl.DTLVardef@1de59d3_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2020

Detecting genetic variation and base modifications together in the same single molecules of DNA and RNA at base pair resolution using a magnetic tweezer platform.

Accurate decoding of nucleic acid variation is important to understand the complexity and regulation of genome function. Here we introduce a single-molecule platform based on magnetic tweezer (MT) technology that can identify and map the positions of sequence variation and multiple base modifications together in the same single molecules of DNA or RNA at single base resolution. Using synthetic templates, we demonstrate that our method can distinguish the most common epigenetic marks on DNA and RNA with high sensitivity, specificity and precision. We also developed a highly specific CRISPR-Cas enrichment strategy to target genomic regions in native DNA without amplification. We then used this method to enrich native DNA from E. coli and characterized the differential levels of adenine and cytosine base modifications together in molecules of up to 5 kb in length. Finally, we enriched the 5 UTR of FMR1 from cells derived from a Fragile X carrier and precisely measured the repeat expansion length and methylation status of each molecule. These results demonstrate that our platform can detect a variety of genetic, epigenetic and base modification changes concomitantly within the same single molecules.

genomics↗