Biology subjects

Brandani, G. B.

Publications and source records attributed to Brandani, G. B..

10 recordsLinked to original sources

bioRxiv2025

Physics-based nucleosome-resolution modeling of epigenetic-driven chromatin domain dynamics

Chromatin spatially organizes the Eukaryotic genome to support key cellular processes such as gene regulation, but the interplay between epigenetics, chromatin structure, and function is still poorly understood. We propose a nucleosome-resolution coarse-grained model that captures the essential features of chromatin organization over multiple scales: from nucleosome dynamics to chromatin fiber folding, and liquid-liquid phase separation. The model describes the effects of DNA linker length, histone tail acetylation, linker histone H1, and multi-bromodomain proteins such as BRD4. It is designed to be experimentally accurate but computationally efficient, allowing the study of 100kb genomic regions on a timescale of seconds with moderate resources. We apply this model to explore the structure and dynamics of two active loci of mouse embryonic stem cells, Pou5f1 and Sox2, as determined by solely their underlying epigenetic patterns. Our simulations reveal that chromatin folds into liquid-like domains characterized by similar histone modifications. These domains are highly dynamic, driving the formation of transient contacts between distant cis-regulatory regions. In silico mutation studies further clarify the distinct roles played by histone acetylation, linker histone, and BRD4. Overall, our physics-based modeling provides evidence that, in addition to other well-established mechanisms, epigenetic-dependent nucleosome-nucleosome interactions can play a key role in shaping the functional organization of genomic loci.

biophysics↗

bioRxiv2025

Multiphase separation in postsynaptic density regulated by membrane geometry via interaction valency and volume

Biomolecular condensates are found at various cellular locations, nucleus, cytoplasm, and membrane. These condensates often contain multiple components and can separate into multiple phases with various morphologies such as core-shell droplets, implicating functional roles. Demixing and arrangements of condensates are determined by competitive interactions and their locations. Recent studies reported a puzzling multiphase morphology in postsynaptic density components: AMPA-receptor, NMDA-receptor, PSD-95, and CaMKII. The multiphase morphology appears reversed when transitioning from the solution to the membrane. Using this system as a model, we study the multiphase behavior of condensates in solution (3D) and domain formation on and beneath the membrane (2D) and elucidate molecular mechanisms behind the puzzle. Our simulations reproduce the core-shell structure in 3D in vitro solution, where AMPA-receptor/PSD-95 form the core and NMDA-receptor/CaMKII form the shell, triggered by CaMKII activation. Then, we obtain a reversed morphology on the membrane. This reversal is primarily driven by CaMKIIs high valency and large volume. We find that, in solution, CaMKIIs non-specific volume interaction dominate, while on the membrane, specific multivalent interactions overcome the excluded volume interaction of CaMKII. The layered structures of receptors and CaMKIIs reduce the excluded volume effects of CaMKII on receptors, making the multivalent interaction dominant. These findings highlight the differences between condensate formation in solution and membrane domain formation, modulated by their layered arrangement. SignificanceMany proteins form condensates in cells, which are often further separated into two or more phases, implicating functional roles. Recent experiments on postsynaptic density proteins provide an interesting puzzle on a multiphasic morphology; while they form core-shell droplets with AMPA-receptor at the core and NMDA receptor in the shell in soluble constructs, the multiphase morphology is reversed when they form nanodomains on the membrane. This study identifies the mechanism behind the puzzle by using computer simulations with a mesoscale model. While active CaMKII, an inducer of the multiphase morphology, has dominant non-specific volume interactions in solution, its specific domain-domain multivalent interaction overcomes the volume interaction under the membrane with their layered arrangement.

biophysics↗

bioRxiv2024

In silico nanoscope to study the interplay of genome organization and transcription regulation

In eukaryotic genomes, regulated access and communication between cis-regulatory elements (CREs) are necessary for enhancer-mediated transcription of genes. The molecular framework of the chromatin organization underlying such communication remains poorly understood. To better understand it, we develop a multiscale modeling pipeline to build near-atomistic models of the 200 kb Nanog gene locus in mouse embryonic stem cells comprising nucleosomes, transcription factors, co-activators, and RNA polymerase II-Mediator complexes. By integrating diverse experimental data, including protein localization, genomic interaction frequencies, cryo-electron microscopy, and single-molecule fluorescence studies, our model offers novel insights into chromatin organization and its role in enhancer-promoter communication. The models equilibrated by high-performance molecular dynamics simulations span a scale of [~]350 nm, revealing an experimentally consistent local and global organization of chromatin and transcriptional machinery. Our models elucidate that the sequence-regulated chromatin accessibility facilitates the recruitment of transcription regulatory proteins exclusively at CREs, guided by the contrasting nucleosome organization compared to other regions. By constructing an experimentally consistent near-atomic model of chromatin in the cellular environment, our approach provides a robust framework for future studies on nuclear compartmentalization, chromatin organization, and transcription regulation.

molecular biology↗

bioRxiv2024

Molecular dynamics simulations of human cohesin subunits identify DNA binding sites and their potential roles in DNA loop extrusion

The SMC complex cohesin mediates interphase chromatin structural formation in eukaryotic cells through DNA loop extrusion. Here, we sought to investigate its mechanism using molecular dynamics simulations. To achieve this, we first constructed the amino-acid-residue-resolution structural models of the cohesin subunits, SMC1, SMC3, STAG1, and NIPBL. By simulating these subunits with double-stranded DNA molecules, we predicted DNA binding patches on each subunit and quantified the affinities of these patches to DNA using their dissociation rate constants as a proxy. Then, we constructed the structural model of the whole cohesin complex and mapped the predicted high-affinity DNA binding patches on the structure. From the spatial relations of the predicted patches, we identified that multiple patches on the SMC1, SMC3, STAG1, and NIPBL subunits form a DNA clamping patch group. The simulations of the whole complex with double-stranded DNA molecules suggest that this patch group facilitates DNA bending and helps capture a DNA segment in the cohesin ring formed by the SMC1 and SMC3 subunits. In previous studies, these have been identified as critical steps in DNA loop extrusion. Therefore, this study provides experimentally testable predictions of DNA binding sites implicated in previously proposed DNA loop extrusion mechanisms and highlights the essential roles of the accessory subunits STAG1 and NIPBL in the mechanism.

molecular biology↗

bioRxiv2024

SMC complex unidirectionally translocates DNA by coupling segment capture with an asymmetric kleisin path

SMC (structural maintenance of chromosomes) protein complexes are ring-shaped molecular motors essential for genome folding. Despite recent progress, the detailed molecular mechanism of DNA translocation in concert with the ATP-driven conformational changes of the complex remains to be clarified. In this study, we elucidated the mechanisms of SMC action on DNA using all-atom and coarse-grained molecular dynamics simulations. We first created a near-atomic full-length model of a prokaryotic SMC-kleisin complex based on experimental structures and implemented ATP-dependent conformational changes using a structure-based coarse-grained model. We further incorporated key protein-DNA hydrogen bond interactions derived from fully atomistic simulations. Extensive simulations of the SMC complex with 800 base pairs of duplex DNA over the ATP cycle observed unidirectional DNA translocation by the SMC complex. The process exhibited a step size of [~]200 base pairs, wherein the SMC complex captured a DNA segment of about the same size within the SMC ring in the engaged state, followed by its pumping into the kleisin ring as ATP was hydrolyzed. Analysis of trajectories identified the asymmetric path of the kleisin as a critical factor for the observed unidirectionality. SIGNIFICANCE STATEMENTRing-shaped SMC (structural maintenance of chromosomes) protein complexes, which are highly conserved across all three domains of life, play an essential role in chromosome organization through a process called DNA loop extrusion. However, the molecular mechanism underlying the ATP-dependent motor activity of SMC complexes remains unclear. Using all-atom and residue-resolution coarse-grained molecular dynamics simulations, we revealed that prokaryotic SMC complexes translocate unidirectionally along DNA via a segment capture mechanism. We found that the unidirectionality arises from the kleisin subunit breaking the symmetry of the ring-shaped SMC complex structure. Our findings provide insights into the molecular motor mechanisms shared by SMC complexes.

biophysics↗

bioRxiv2023

Multiscale Bayesian Simulations Reveal Functional Chromatin Condensation of Gene Loci

Chromatin, the complex assembly of DNA and associated proteins, plays a pivotal role in orchestrating various genomic functions. To aid our understanding of the principles underlying chromatin organization, we introduce Hi-C metainference, a Bayesian approach that integrates Hi-C contact frequencies into multiscale prior models of chromatin. This approach combines both bottom-up (the physics-based prior) and top-down (the data-driven posterior) strategies to characterize the 3D organization of a target genomic locus. We first demonstrate the capability of this method to accurately reconstruct the structural ensemble and the dynamics of a system from contact information. We then apply the approach to investigate the Sox2, Pou5f1, and Nanog loci of mouse embryonic stem cells using a bottom-up chromatin model at 1kb resolution. We observe that the studied loci are conformationally heterogeneous and organized as crumpled globules, favoring contacts between distant enhancers and promoters. Using nucleosome-resolution simulations, we then reveal how the Nanog gene is functionally organized across the multiple scales of chromatin. At the local level, diverse chromatin folding motifs correlate with epigenetics, with open chromatin predominantly observed at cis-regulatory elements and compact tetranucleosomes in between. At the larger scale, we find that enhancer-promoter contacts are driven by the transient condensation of chromatin into compact domains stabilized by extensive inter-nucleosome interactions. Overall, this work highlights the condensed, but dynamic nature of chromatin in vivo, contributing to a deeper understanding of gene structure-function relationships.

biophysics↗

bioRxiv2021

DNA translocase repositions a nucleosome by the lane-switch mechanism

Translocases such as DNA/RNA polymerases, replicative helicases, and exonucleases are involved in eukaryotic DNA transcription, replication, and repair. Since eukaryotic genomic DNA wraps around histone core complexes and forms nucleosomes, translocases inevitably encounter nucleosomes. Previous studies have shown that a histone core complex repositions upstream (downstream) when SP6RNA or T7 RNA polymerase (bacterial exonuclease, RecBCD) partially unwraps nucleosomal DNA. However, the molecular mechanism of the downstream repositioning remains unclear. In this study, we identify the lane-shift mechanism for downstream nucleosome repositioning via coarse-grained molecular dynamics simulations, which we validated by restriction enzyme digestion assays and deep sequencing assays. In this mechanism, after a translocase unwraps nucleosomal DNA up to the site proximal to the dyad, the remaining wrapped DNA switches its binding region (lane) to that vacated by the unwrapping, and the downstream DNA rewraps, completing downstream repositioning. This mechanism may have crucial implications for transcription through nucleosomes, histone recycling, and nucleosome remodeling. SIGNIFICANCEEukaryotic chromosomes are composed of repeating subunits termed nucleosomes. Thus, proteins that translocate along the chromosome, DNA translocases, inevitably collide with nucleosomes. Previous studies revealed that a translocase repositions a nucleosome upstream or downstream upon their collision. Though the molecular mechanisms of the upstream repositioning have been extensively studied, that of downstream repositioning remains elusive. In this study, we performed coarse-grained molecular dynamics simulations, proposed the lane-shift mechanism for downstream repositioning, and validated this mechanism by restriction enzyme digestion assays and deep sequencing assays. This mechanism has broad implications for how translocases deal with nucleosomes for their functions.

biophysics↗

bioRxiv2021

The Kinetic Landscape of Nucleosome Assembly: A Coarse-Grained Molecular Dynamics Study

The organization of nucleosomes along the Eukaryotic genome is maintained over time despite disruptive events such as replication. During this complex process, histones and DNA can form a variety of non-canonical nucleosome conformations, but their precise molecular details and roles during nucleosome assembly remain unclear. In this study, employing coarse-grained molecular dynamics simulations and Markov state modeling, we characterized the complete kinetics of nucleosome assembly. On the nucleosome-positioning 601 DNA sequence, we observe a rich transition network among various canonical and non-canonical tetrasome, hexasome, and nucleosome conformations. A low salt environment makes nucleosomes stable, but the kinetic landscape becomes more rugged, so that the system is more likely to be trapped in off-pathway partially assembled intermediates. We find that the co-operativity between DNA bending and histone association enables positioning sequence motifs to direct the assembly process, with potential implications for the dynamic organization of nucleosomes on real genomic sequences.

biophysics↗

bioRxiv2018

Chromatin remodelers couple inchworm motion with twist-defect formation to slide nucleosomal DNA

ATP-dependent chromatin remodelers control genome organization by repositioning, ejecting, or editing nucleosomes, activities that confer them essential regulatory roles on gene expression and DNA replication. Here, we investigate the mechanism of active DNA sliding by remodelers, which underlies most remodeling functions, via molecular dynamics simulations of the Snf2 remodeler in complex with a nucleosome. During its inchworm motion driven by ATP consumption, the remodeler overwrites the original nucleosome energy landscape via steric and electrostatic interactions to induce sliding of nucleosomal DNA. Repositioning is initiated at the remodeler binding location via the generation of twist defects, which then spontaneously propagate to complete sliding throughout the entire nucleosome. We also reveal how remodeler mutations and DNA sequence control active nucleosome repositioning. These results offer a mechanistic understanding of chromatin remodeling important for the interpretation of experimental studies.

biophysics↗

bioRxiv2017

Sequence-dependent nucleosome sliding in rotation-coupled and uncoupled modes revealed by molecular simulations

While nucleosome positioning on eukaryotic genome play important roles for genetic regulation, molecular mechanisms of nucleosome positioning and sliding along DNA are not well understood. Here we investigated thermally-activated spontaneous nucleosome sliding mechanisms developing and applying a coarse-grained molecular simulation method that incorporates both long-range electrostatic and short-range hydrogen-bond interactions between histone octamer and DNA. The simulations revealed two distinct sliding modes depending on the nucleosomal DNA sequence. A uniform DNA sequence showed frequent sliding with one base pair step in a rotation-coupled manner, akin to screw-like motions. On the contrary, a strong positioning sequence, the so-called 601 sequence, exhibits rare, abrupt transitions of five and ten base pair steps without rotation. Moreover, we evaluated the importance of hydrogen bond interactions on the sliding mode, finding that strong and weak bonds favor respectively the rotation-coupled and -uncoupled sliding movements.\n\nAuthor summaryNucleosomes are fundamental units of chromatin folding consisting of double-stranded DNA wrapped [~]1.7 times around a histone octamer. By densely populating the eukaryotic genome, nucleosomes enable efficient genome compaction inside the cellular nucleus. However, the portion of DNA occupied by a nucleosome can hardly be accessed by other DNA-binding proteins, obstructing fundamental cellular processes such as DNA replication and transcription. DNA compaction and access by other proteins can simultaneously be achieved via the dynamical repositioning of nucleosomes, which can slide along the DNA sequence. In this study, we developed and used coarse-grained molecular dynamics simulations to reveal the molecular details of nucleosome sliding. We find that the sliding mode is highly dependent on the underlying DNA sequence. Specifically, a sequence with a strong nucleosome positioning signal slides via large jumps by five and ten base pairs, preserving the optimal DNA bending profile. On the other hand, uniform sequences without the positioning signal slide via a screw-like motion of DNA, one base pair at the time. These results show that sequence has a large effect not only on the formation of nucleosomes, but also on the kinetics of repositioning.

biophysics↗