Biology subjects

Watkins, S. L.

Publications and source records attributed to Watkins, S. L..

3 recordsLinked to original sources

bioRxiv2026

Decoding the RNA Splicing Network in HNRNPH2-R114W Brain Organoids

Autism spectrum disorder and related neurodevelopmental diseases are increasingly linked to disrupted RNA processing during corticogenesis, yet resolving variant-specific mechanisms during early human development remains challenging. We generated isogenic cerebral organoids from a patient iPSC line carrying the HNRNPH2-R114W mutation and its CRISPR-corrected counterpart. Developmental validation by time-course imaging, followed by bulk RNA-Seq at day 20, showed comparable early differentiation and corticogenesis. Genotype was the primary determinant of global expression state, with organized remodeling of neuronal and metabolic programs. Splicing analyses demonstrated junction-dominant dysregulation, where 6,310 junctions exhibited significant differential usage. Junction-level resolution revealed a dominant genotype-related signal than exon-centric analysis. A structural docking model shows HNRNPH2 RNA recognition motifs 1-3 within a spliceosomal context and position residue R114 at an RNA-contact cleft, consistent with direct disruption of RNA engagement by R114W. Integrating differential expression with event- and isoform-level evidence prioritized convergent regulatory hubs enriched for axon guidance, synaptic signaling, and extracellular matrix programs. Single-cell RNA-Seq deconvolution revealed modest compositional shifts that do not account for the magnitude of transcriptional and splicing alterations. Collectively, these data support that HNRNPH2-R114W drives pervasive, junction-centered RNA rewiring in human cortical organoids and defines isoform-level endpoints for mechanistic studies and therapeutic testing in an isogenic developmental model.

neuroscience↗

bioRxiv2025

Molecular Dynamics Reveal Base Flipping as a Key Mechanism in Tc3a Transposase DNA Recognition

Transposons, or "jumping genes," are mobile genetic elements that reshape genomes. The Tc1/Mariner family, including Tc3a transposase, has been studied for decades, but key aspects of its DNA recognition and cleavage remain unresolved. Existing models propose a homo-dimeric process for DNA excision and integration, yet conflicting hypotheses exist about the initial steps of DNA cleavage and target site recognition. Here, we reveal a previously unrecognized base-flipping mechanism in Tc3a transposase that challenges long-standing models of Mariner transposon activity. Using molecular dynamics simulations, we demonstrate that N-terminal acetylation induces a structural shift in the transposase-DNA complex, forcing a nucleotide in the inverted repeat to flip outward. This base-flipping event alters the local DNA conformation, creating torsional strain that may facilitate transposon inverted terminal repeat association. Unlike prior models, which assumed the entire protein, or protein dimers, were needed to illicit DNA interactions necessary for any types of torsion, our findings indicate that transposase bipartite binding itself actively reshapes DNA structure. We further show that this mechanism is dependent on the bipartite helix-turn-helix domains, which differentially contribute to DNA stabilization and recognition. Our simulations also reveal that the processed N-terminus plays an unexpected role in modulating DNA binding affinity, contradicting previous assumptions that it was structurally inconsequential. Together, the two helix-turn-helix motifs act to propagate the force caused by base flipping along one direction of the cognate DNA double helix axis. The fact that previous structural studies lacked this amino acid modification may explain why this mechanism was overlooked. These results provide a new framework for understanding transposase-DNA interactions, highlighting how a single molecular modification can trigger major DNA rearrangements. This discovery not only redefines the Tc3a transposition process but also calls for a reassessment of long-standing Mariner family transposon models.

biochemistry↗

bioRxiv2021

Molecular Dynamics Simulations of Phytopthora sojae Avirulence Factor 5, PsAvh5, Define Membrane Binding and Inositolphosphate-3'-phosphate Interactions and Protein Mechanics

Phytopthora Avirulence proteins are a primary target for development of rational chemical and biological control of some of the most devastating plant pathogens. Despite the sequencing of entire genomes, and characterization of many of these proteins at the chemical level, many questions remain regarding actual chemical and biological interactions involved. In addition, disputed roles of ligands, such as Inositolphosphate-3-phosphate and amino acids of important function remain unclear. To address some of these issues, we developed molecular models from structural elements and published data for Phytopthora sojae avirulence protein 5. Molecular dynamics simulations are used to study protein function, interactions involved primarily with lipids and membranes, and inositol derivatives. Our findings indicate that the protein is stable as a monomer, and in a dimeric form. Also, that these proteins interact with Inositolphosphate-3phosphate as a necessary membrane element, in binding. We identified several amino acids of importance, additional to defining the mechanical features of the protein within the binding process to different membranes. A high affinity, comparable to other membrane surface binding molecules of -219.54 Kcal for the dimer, and -176.61 for the monomer were determined. With either form, we found the inositolphosphate-3-phosphate to be essential in the membrane binding process. Our findings answer some of the debated questions while creating a point to further test avirulence proteins in general for functional aspects. Additionally, the structures and data can be utilized to provide a better starting point for rational design approaches to control this pathogen.

biophysics↗