Search bioRxiv⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2025.04.09.648031

DNA six-way junction conformations and their use in a 2D square lattice

Baten, A. J.·Mason, H. G.·Veneziano, R.·Bricker, W. P.
View original source ↗Export metadata ↓

Abstract

In this paper, we investigate the conformational landscape of the DNA six-way junction (6WJ), a higher-order extension of the DNA four-way junction (4WJ), to de-termine preferred structural isomers. The 6WJ allows for unique junction-level topolo-gies, and could be used to create novel DNA nanostructures, but the conformational landscape for the 6WJ is much more complex than the 4WJ. Our proposed confor-mational landscape for the 6WJ includes eight unique structural motifs, including an unstacked motif, five distinct stacked motifs, and two twisted motifs, and we estimate that there are forty structural isomers for the 6WJ, compared to only three for the 4WJ. To gain insight into these conformations, we perform all-atom molecular dynam-ics (MD) simulations on fourteen of the structural isomers. Our analysis shows that each 6WJ motif can be distinguished by its duplex stacking angles, and only a few of the 6WJ isomers have favorable free energies, which include the planar parallel isomers, the twisted isomers, and one of the orthogonal isomers. Lastly, to confirm that these stacked 6WJ motifs could be used to create larger-scale DNA nanostructures, we design a 2D square lattice using four orthogonal motifs connected in a 2x2 arrangement. Experimental characterization of this 2D lattice shows folding into the correct size and shape, further confirmed by AFM images of the DNA tiles, and an MD simulation that shows a slight twist of the lattice structure from orthogonality. This is the first 2D tile assembly built using an orthogonal junction-level topology for DNA nanotechnology, allowing a smaller mesh size than is possible with the 4WJ architecture.

Explore related subjects

bioengineeringPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Baten, A. J., Mason, H. G., Veneziano, R., Bricker, W. P.. 2025-04-15. DNA six-way junction conformations and their use in a 2D square lattice. https://doi.org/10.1101/2025.04.09.648031

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Selected for the surface: adaptive laboratory evolution for accelerated cathodic biofilm development in Cupriavidus necator

Cupriavidus necator is a metabolically versatile facultative chemolithoautotroph and a promising chassis for sustainable bioproduction from CO2. Efficient coupling of its metabolism to renewable electricity through microbial electrosynthesis (MES) requires the organism to form stable, electroactive biofilms on cathodes. This step is hindered under oxic conditions (oxic MES, oMES) by electrochemically generated reactive oxygen species (ROS), mass-transfer limitations and poor biofilm stability. Systematic strategies to improve C. necator biofilm formation on cathodes have been lacking. Here we used adaptive laboratory evolution (ALE) to address this gap. The progenitor strain colonised graphite cathodes only after an initial heterotrophic phase and formed spatially uneven biofilms whose distribution mirrored the local flow field, as resolved by micro-particle tracking velocimetry. Populations were then selected over five passages under controlled, oxygen-limited cathodic conditions. Resequencing revealed that the recovered population was genetically heterogeneous, and its dominant isolate, strain 890, was characterised in a microfluidic oMES platform. Strain 890 reached the target electrode coverage in about one day instead of six and maintained a robust biofilm at the progenitor's steady-state biovolume, whereas the multi-strain pool peaked higher but proved unstable. Relative to the progenitor, the accelerated biofilm phenotype of strain 890 is driven by a single novel mutation in a bifunctional diguanylate cyclase/phosphodiesterase locus, which likely encodes a central regulator of the planktonic-to-biofilm transition. The steady-state biovolume itself remained bounded, most probably due to a mass-transfer ceiling set by the counter-directional supply of H2 and O2. This work shows that ALE renders the initial-attachment barrier of cathodic biofilm formation genetically tractable, providing strain 890 as a robust chassis and a well-defined starting point for oMES-based CO2 valorisation.

bioengineering↗
bioRxiv2026

Engineering a High-Fidelity CRISPR-Cas13d System for Targeted RNA Depletion in Plasmodium falciparum

Functional genomics in Plasmodium falciparum is severely constrained by the evolutionary absence of an endogenous RNA interference pathway and the requirement for time-consuming direct target locus modification by conventional conditional genetic engineering systems. To overcome these limitations, we established a programmable post-transcriptional gene silencing system in P. falciparum using High-Fidelity RfxCas13d (HF-RfxCas13d), an RNA-guided RNA nuclease that has been previously tested in several eukaryotic hosts. Using CRISPR-SpyCas9 we integrated a HF-RfxCas13d expression cassette at the dispensable pfs47 locus and generated a stable marker-free transgenic line capable of constitutive HF-RfxCas13d cytosolic expression without impairing intraerythrocytic asexual growth. Episomal delivery of single or dual guide RNAs (gRNAs) achieved significant target mRNA depletion (41%-76%) across three endogenous loci atp4, ap2-exp, ap2-g genes. Applying this system to the master transcriptional regulator of gametocytogenesis (ap2-g) resulted in transcript depletion that functionally impaired sexual differentiation in committed parasites, reducing significantly the Sexual Conversion Rate a 70%. Overall, our novel HF-Cas13d line provides a flexible, scalable, and non-locus-modifying addition to the P. falciparum genetic toolbox to accelerate functional genomics and malaria drug target identification.

bioengineering↗
bioRxiv2026

DECODE: DEcomposition of COupled and DEcoupled Brain States: Separating Structure-Coupled and Structure-Decoupled Functional Organization in Alzheimers Disease

Brain functional activity emerges within an underlying structural scaffold. While structural network connectivity provides a relatively stable anatomical backbone for large-scale communication, functional interactions are high-dimensional and temporally dynamic, forming recurrent patterns that can be represented in a latent space of transient functional states. Yet, how these transient functional states are organized and the extent to which they remain constrained by structural network connectivity or become relatively decoupled from structural organization over time remain poorly understood. To address this gap, we introduce DECODE (DEcomposition of COupled and DEcoupled Brain States), a data-driven latent-space framework that uses independent component analysis (ICA) and multimodal data to decompose resting-state dynamic functional network connectivity into a lower-dimensional representation of transient functional states. These states capture temporally varying structure-coupled and structure-decoupled patterns using diffusion MRI-derived structural network connectivity priors. Structure-coupled states reflect functional configurations more strongly anchored to major structural motifs, whereas structure-decoupled states represent configurations less constrained by the identified structural organization, while potentially remaining within structurally defined dynamical bounds. DECODE identifies reproducible structure-coupled and structure-decoupled states, with mean squared error reconstruction of dynamic information < 0.1 and adjusted R-squared = 0.4. Applied to Alzheimers disease (AD), DECODE revealed stronger group differences (FDR-corrected p-value 0.05) in both connectivity organization and temporal dynamics for decoupled states. These findings suggest that functional states less constrained by identified structural organization may be more vulnerable to AD-related disruption, whereas structure-coupled states may be relatively preserved. More broadly, DECODE provides a reproducible and generalizable framework for quantifying how functional dynamics are shaped by, or diverge from, structural organization by incorporating a latent representation of the structural connectome as an informative prior. Our findings established structural and functional alignment variation by distinguishing dynamic functional patterns closely coupled with structure from those that deviate from it.

bioengineering↗