Weak Pinning and Long-Range Anticorrelated Motion of Phase Boundaries in Driven Diffusive Systems.

We show that domain walls separating coexisting extremal current phases in driven diffusive systems exhibit complex stochastic dynamics with a subdiffusive temporal growth of position fluctuations due to long-range anticorrelated current fluctuations and a weak pinning at long times. This weak pinning manifests itself in a saturated width of the domain wall position fluctuations that increases sublinearly with the system size. As a function of time t and system size L, the width w(t,L) has a scaling behavior w(t,L)=L^{3/4}f(t/L^{9/4}), with f(u) constant for u≫1 and f(u)∼u^{1/3} for u≪1. An Orstein-Uhlenbeck process with long-range anticorrelated noise is shown to capture this scaling behavior. The exponent 9/4 is a new dynamical exponent for relaxation processes in driven diffusive systems.

DNA double-strand break-capturing nuclear envelope tubules drive DNA repair.

Current models suggest that DNA double-strand breaks (DSBs) can move to the nuclear periphery for repair. It is unclear to what extent human DSBs display such repositioning. Here we show that the human nuclear envelope localizes to DSBs in a manner depending on DNA damage response (DDR) kinases and cytoplasmic microtubules acetylated by α-tubulin acetyltransferase-1 (ATAT1). These factors collaborate with the linker of nucleoskeleton and cytoskeleton complex (LINC), nuclear pore complex (NPC) protein NUP153, nuclear lamina and kinesins KIF5B and KIF13B to generate DSB-capturing nuclear envelope tubules (dsbNETs). dsbNETs are partly supported by nuclear actin filaments and the circadian factor PER1 and reversed by kinesin KIFC3. Although dsbNETs promote repair and survival, they are also co-opted during poly(ADP-ribose) polymerase (PARP) inhibition to restrain BRCA1-deficient breast cancer cells and are hyper-induced in cells expressing the aging-linked lamin A mutant progerin. In summary, our results advance understanding of nuclear structure-function relationships, uncover a nuclear-cytoplasmic DDR and identify dsbNETs as critical factors in genome organization and stability.

High-throughput functional analysis of regulatory variants using a massively parallel reporter assay.

Association studies describe genetic associations between noncoding variants and disease susceptibility; however, they do not provide functional insight into the underlying molecular mechanisms of these variants. We present a protocol to assay the regulatory potential of thousands of noncoding variants using massively parallel reporter assays. We describe steps for oligo design, generating a plasmid pool, and extracting tag-seq libraries from cells to quantify the tested sequences. For complete details on the use and execution of this protocol, please refer to Oliveros and Delfosse et al.1.

Overcrowding induces fast colloidal solitons in a slowly rotating potential landscape.

Collective particle transport across periodic energy landscapes is ubiquitously present in many condensed matter systems spanning from vortices in high-temperature superconductors, frictional atomic sliding, driven skyrmions to biological and active matter. Here we report the emergence of fast solitons propagating against a rotating optical landscape. These experimentally observed solitons are stable cluster waves that originate from a coordinated particle exchange process which occurs when the number of trapped microparticles exceeds the number of potential wells. The size and speed of individual solitons rapidly increase with the particle diameter as predicted by theory and confirmed by numerical simulations. We show that when several solitons coexist, an effective repulsive interaction can stabilize their propagation along the periodic potential. Our experiments demonstrate a generic mechanism for cluster-mediated transport with potential applications to condensed matter systems on different length scales.

Single-file transport of binary hard-sphere mixtures through periodic potentials.

Single-file transport occurs in various scientific fields, including diffusion through nanopores, nanofluidic devices, and cellular processes. We here investigate the impact of polydispersity on particle currents for single-file Brownian motion of hard spheres when they are driven through periodic potentials by a constant drag force. Through theoretical analysis and extensive Brownian dynamics simulations, we unveil the behavior of particle currents for random binary mixtures. The particle currents show a recurring pattern in dependence of the hard-sphere diameters and mixing ratio. We explain this recurrent behavior by showing that a basic unit cell exists in the space of the two hard-sphere diameters. Once the behavior of an observable inside the unit cell is determined, it can be inferred for any diameter. The overall variation of particle currents with the mixing ratio and hard-sphere diameters is reflected by their variation in the limit where the system is fully covered by hard spheres. In this limit, the currents can be predicted analytically. Our analysis explains the occurrence of pronounced maxima and minima of the currents by changes in the effective potential barrier for the center-of-mass motion.

Signatures of geostrophic turbulence in power spectra and third-order structure function of offshore wind speed fluctuations.

We analyze offshore wind speeds with a time resolution of one second over a long period of 20 months for different heights above the sea level. Energy spectra extending over more than seven decades give a comprehensive picture of wind fluctuations, including intermittency effects at small length scales and synoptic weather phenomena at large scales. The spectra S(f) show a scaling behavior consistent with three-dimensional turbulence at high frequencies f, followed by a regime at lower frequencies, where fS(f) varies weakly. Lowering the frequency below a crossover frequency [Formula: see text], a rapid rise of fS(f) occurs. An analysis of the third-order structure function [Formula: see text] of wind speed differences for a given time lag [Formula: see text] shows a rapid change from negative to positive values of [Formula: see text] at [Formula: see text]. Remarkably, after applying Taylor's hypothesis locally, we find the third-order structure function to exhibit a behavior very similar to that obtained previously from aircraft measurements at much higher altitudes in the atmosphere. In particular, the third-order structure function grows linearly with the separation distance for negative [Formula: see text], and with the third power for positive [Formula: see text]. This allows us to estimate energy and enstrophy dissipation rates for offshore wind. The crossover from negative to positive values occurs at about the same separation distance of 400 km as found from the aircraft measurements, suggesting that this length is independent of the altitude in the atmosphere.

Systematic characterization of regulatory variants of blood pressure genes.

High blood pressure (BP) is the major risk factor for cardiovascular disease. Genome-wide association studies have identified genetic variants for BP, but functional insights into causality and related molecular mechanisms lag behind. We functionally characterize 4,608 genetic variants in linkage with 135 BP loci in vascular smooth muscle cells and cardiomyocytes by massively parallel reporter assays. High densities of regulatory variants at BP loci (i.e., ULK4, MAP4, CFDP1, PDE5A) indicate that multiple variants drive genetic association. Regulatory variants are enriched in repeats, alter cardiovascular-related transcription factor motifs, and spatially converge with genes controlling specific cardiovascular pathways. Using heuristic scoring, we define likely causal variants, and CRISPR prime editing finally determines causal variants for KCNK9, SFXN2, and PCGF6, which are candidates for developing high BP. Our systems-level approach provides a catalog of functionally relevant variants and their genomic architecture in two trait-relevant cell lines for a better understanding of BP gene regulation.

Scaling laws for single-file diffusion of adhesive particles.

Single-file diffusion refers to the Brownian motion in narrow channels where particles cannot pass each other. In such processes, the diffusion of a tagged particle is typically normal at short times and becomes subdiffusive at long times. For hard-sphere interparticle interaction, the time-dependent mean squared displacement of a tracer is well understood. Here we develop a scaling theory for adhesive particles. It provides a full description of the time-dependent diffusive behavior with a scaling function that depends on an effective strength of adhesive interaction. Particle clustering induced by the adhesive interaction slows down the diffusion at short times, while it enhances subdiffusion at long times. The enhancement effect can be quantified in measurements irrespective of how tagged particles are injected into the system. Combined effects of pore structure and particle adhesiveness should speed up translocation of molecules through narrow pores.

Brownian dynamics simulations of hard rods in external fields and with contact interactions.

We propose a simulation method for Brownian dynamics of hard rods in one dimension for arbitrary continuous external force fields. It is an event-driven procedure based on the fragmentation and mergers of clusters formed by particles in contact. It allows one to treat particle interactions in addition to the hard-sphere exclusion as long as the corresponding interaction forces are continuous functions of the particle coordinates. We furthermore develop a treatment of sticky hard spheres as described by Baxter's contact interaction potential.

Hydrodynamic interactions hinder transport of flow-driven colloidal particles.

The flow-driven transport of interacting micron-sized particles occurs in many soft matter systems spanning from the translocation of proteins to moving emulsions in microfluidic devices. Here we combine experiments and theory to investigate the collective transport properties of colloidal particles along a rotating ring of optical traps. In the corotating reference frame, the particles are driven by a vortex flow of the surrounding fluid. When increasing the depth of the optical potential, we observe a jamming behavior that manifests itself in a strong reduction of the current with increasing particle density. We show that this jamming is caused by hydrodynamic interactions that enhance the energetic barriers between the optical traps. This leads to a transition from an over- to an under-critical tilting of the potential in the corotating frame. Based on analytical considerations, the enhancement effect is estimated to increase with increasing particle size or decreasing radius of the ring of traps. Measurements for different ring radii and Stokesian dynamics simulations for corresponding particle sizes confirm this. The enhancement of potential barriers in the flow-driven system is contrasted to the reduction of barriers in a force-driven one. This diverse behavior demonstrates that hydrodynamic interactions can have a very different impact on the collective dynamics of many-body systems. Applications to soft matter and biological systems require careful consideration of the driving mechanism and of the role of hydrodynamic interactions.

Identifying Tissue- and Cohort-Specific RNA Regulatory Modules in Cancer Cells Using Multitask Learning.

MicroRNA (miRNA) alterations significantly impact the formation and progression of human cancers. miRNAs interact with messenger RNAs (mRNAs) to facilitate degradation or translational repression. Thus, identifying miRNA-mRNA regulatory modules in cohorts of primary tumor tissues are fundamental for understanding the biology of tumor heterogeneity and precise diagnosis and treatment. We established a multitask learning sparse regularized factor regression (MSRFR) method to determine key tissue- and cohort-specific miRNA-mRNA regulatory modules from expression profiles of tumors. MSRFR simultaneously models the sparse relationship between miRNAs and mRNAs and extracts tissue- and cohort-specific miRNA-mRNA regulatory modules separately. We tested the model's ability to determine cohort-specific regulatory modules of multiple cancer cohorts from the same tissue and their underlying tissue-specific regulatory modules by extracting similarities between cancer cohorts (i.e., blood, kidney, and lung). We also detected tissue-specific and cohort-specific signatures in the corresponding regulatory modules by comparing our findings from various other tissues. We show that MSRFR effectively determines cancer-related miRNAs in cohort-specific regulatory modules, distinguishes tissue- and cohort-specific regulatory modules from each other, and extracts tissue-specific information from different cohorts of disease-related tissue. Our findings indicate that the MSRFR model can support current efforts in precision medicine to define tumor-specific miRNA-mRNA signatures.

Mutant Phosphodiesterase 3A Protects From Hypertension-Induced Cardiac Damage.

BACKGROUND: Phosphodiesterase 3A (PDE3A) gain-of-function mutations cause hypertension with brachydactyly (HTNB) and lead to stroke. Increased peripheral vascular resistance, rather than salt retention, is responsible. It is surprising that the few patients with HTNB examined so far did not develop cardiac hypertrophy or heart failure. We hypothesized that, in the heart, PDE3A mutations could be protective. METHODS: We studied new patients. CRISPR-Cas9-engineered rat HTNB models were phenotyped by telemetric blood pressure measurements, echocardiography, microcomputed tomography, RNA-sequencing, and single nuclei RNA-sequencing. Human induced pluripotent stem cells carrying PDE3A mutations were established, differentiated to cardiomyocytes, and analyzed by Ca2+ imaging. We used Förster resonance energy transfer and biochemical assays. RESULTS: We identified a new PDE3A mutation in a family with HTNB. It maps to exon 13 encoding the enzyme's catalytic domain. All hitherto identified HTNB PDE3A mutations cluster in exon 4 encoding a region N-terminally from the catalytic domain of the enzyme. The mutations were recapitulated in rat models. Both exon 4 and 13 mutations led to aberrant phosphorylation, hyperactivity, and increased PDE3A enzyme self-assembly. The left ventricles of our patients with HTNB and the rat models were normal despite preexisting hypertension. A catecholamine challenge elicited cardiac hypertrophy in HTNB rats only to the level of wild-type rats and improved the contractility of the mutant hearts, compared with wild-type rats. The ß-adrenergic system, phosphodiesterase activity, and cAMP levels in the mutant hearts resembled wild-type hearts, whereas phospholamban phosphorylation was decreased in the mutants. In our induced pluripotent stem cell cardiomyocyte models, the PDE3A mutations caused adaptive changes of Ca2+ cycling. RNA-sequencing and single nuclei RNA-sequencing identified differences in mRNA expression between wild-type and mutants, affecting, among others, metabolism and protein folding. CONCLUSIONS: Although in vascular smooth muscle, PDE3A mutations cause hypertension, they confer protection against hypertension-induced cardiac damage in hearts. Nonselective PDE3A inhibition is a final, short-term option in heart failure treatment to increase cardiac cAMP and improve contractility. Our data argue that mimicking the effect of PDE3A mutations in the heart rather than nonselective PDE3 inhibition is cardioprotective in the long term. Our findings could facilitate the search for new treatments to prevent hypertension-induced cardiac damage.

A noncoding single-nucleotide polymorphism at 8q24 drives IDH1-mutant glioma formation.

Establishing causal links between inherited polymorphisms and cancer risk is challenging. Here, we focus on the single-nucleotide polymorphism rs55705857, which confers a sixfold greater risk of isocitrate dehydrogenase (IDH)-mutant low-grade glioma (LGG). We reveal that rs55705857 itself is the causal variant and is associated with molecular pathways that drive LGG. Mechanistically, we show that rs55705857 resides within a brain-specific enhancer, where the risk allele disrupts OCT2/4 binding, allowing increased interaction with the Myc promoter and increased Myc expression. Mutating the orthologous mouse rs55705857 locus accelerated tumor development in an Idh1R132H-driven LGG mouse model from 472 to 172 days and increased penetrance from 30% to 75%. Our work reveals mechanisms of the heritable predisposition to lethal glioma in ~40% of LGG patients.

Solitons in Overdamped Brownian Dynamics.

Solitons are commonly known as waves that propagate without dispersion. Here, we show that they can occur for driven overdamped Brownian dynamics of hard spheres in periodic potentials at high densities. The solitons manifest themselves as periodic sequences of different assemblies of particles moving in the limit of zero noise, where transport of single particles is not possible. They give rise to particle currents at even low temperature that appear in bandlike structures around certain hard-sphere diameters. At high temperatures, the bandlike structures are washed out by the noise, but the particle transport is still dominated by the solitons. All these predicted features should occur in a broad class of periodic systems and are amenable to experimental tests.

Hypomorphic GINS3 variants alter DNA replication and cause Meier-Gorlin syndrome.

The eukaryotic CDC45/MCM2-7/GINS (CMG) helicase unwinds the DNA double helix during DNA replication. The GINS subcomplex is required for helicase activity and is, therefore, essential for DNA replication and cell viability. Here, we report the identification of 7 individuals from 5 unrelated families presenting with a Meier-Gorlin syndrome-like (MGS-like) phenotype associated with hypomorphic variants of GINS3, a gene not previously associated with this syndrome. We found that MGS-associated GINS3 variants affecting aspartic acid 24 (D24) compromised cell proliferation and caused accumulation of cells in S phase. These variants shortened the protein half-life, altered key protein interactions at the replisome, and negatively influenced DNA replication fork progression. Yeast expressing MGS-associated variants of PSF3 (the yeast GINS3 ortholog) also displayed impaired growth, S phase progression defects, and decreased Psf3 protein stability. We further showed that mouse embryos homozygous for a D24 variant presented intrauterine growth retardation and did not survive to birth, and that fibroblasts derived from these embryos displayed accelerated cellular senescence. Taken together, our findings implicate GINS3 in the pathogenesis of MGS and support the notion that hypomorphic variants identified in this gene impaired cell and organismal growth by compromising DNA replication.

Whole genome sequencing delineates regulatory, copy number, and cryptic splice variants in early onset cardiomyopathy.

Cardiomyopathy (CMP) is a heritable disorder. Over 50% of cases are gene-elusive on clinical gene panel testing. The contribution of variants in non-coding DNA elements that result in cryptic splicing and regulate gene expression has not been explored. We analyzed whole-genome sequencing (WGS) data in a discovery cohort of 209 pediatric CMP patients and 1953 independent replication genomes and exomes. We searched for protein-coding variants, and non-coding variants predicted to affect the function or expression of genes. Thirty-nine percent of cases harbored pathogenic coding variants in known CMP genes, and 5% harbored high-risk loss-of-function (LoF) variants in additional candidate CMP genes. Fifteen percent harbored high-risk regulatory variants in promoters and enhancers of CMP genes (odds ratio 2.25, p = 6.70 × 10-7 versus controls). Genes involved in α-dystroglycan glycosylation (FKTN, DTNA) and desmosomal signaling (DSC2, DSG2) were most highly enriched for regulatory variants (odds ratio 6.7-58.1). Functional effects were confirmed in patient myocardium and reporter assays in human cardiomyocytes, and in zebrafish CRISPR knockouts. We provide strong evidence for the genomic contribution of functionally active variants in new genes and in regulatory elements of known CMP genes to early onset CMP.

Systematic mapping of nuclear domain-associated transcripts reveals speckles and lamina as hubs of functionally distinct retained introns.

The nucleus is highly compartmentalized through the formation of distinct classes of membraneless domains. However, the composition and function of many of these structures are not well understood. Using APEX2-mediated proximity labeling and RNA sequencing, we surveyed human transcripts associated with nuclear speckles, several additional domains, and the lamina. Remarkably, speckles and lamina are associated with distinct classes of retained introns enriched in genes that function in RNA processing, translation, and the cell cycle, among other processes. In contrast to the lamina-proximal introns, retained introns associated with speckles are relatively short, GC-rich, and enriched for functional sites of RNA-binding proteins that are concentrated in these domains. They are also highly differentially regulated across diverse cellular contexts, including the cell cycle. Thus, our study provides a resource of nuclear domain-associated transcripts and further reveals speckles and lamina as hubs of distinct populations of retained introns linked to gene regulation and cell cycle progression.

Hydrodynamic Interactions Can Induce Jamming in Flow-Driven Systems.

Hydrodynamic interactions between fluid-dispersed particles are ubiquitous in soft matter and biological systems and they give rise to intriguing collective phenomena. While it was reported that these interactions can facilitate force-driven particle motion over energetic barriers, here we show the opposite effect in a flow-driven system, i.e., that hydrodynamic interactions hinder transport across barriers. We demonstrate this result by combining experiments and theory. In the experiments, we drive colloidal particles using rotating optical traps, thus creating a vortex flow in the corotating reference frame. We observe a jamminglike decrease of particle currents with density for large barriers between traps. The theoretical model shows that this jamming arises from hydrodynamic interactions between the particles. The impact of hydrodynamic interactions is reversed compared to force-driven motion, suggesting that our findings are a generic feature of flow-driven transport.

Genome-wide CRISPR interference screen identifies long non-coding RNA loci required for differentiation and pluripotency.

Although many long non-coding RNAs (lncRNAs) exhibit lineage-specific expression, the vast majority remain functionally uncharacterized in the context of development. Here, we report the first described human embryonic stem cell (hESC) lines to repress (CRISPRi) or activate (CRISPRa) transcription during differentiation into all three germ layers, facilitating the modulation of lncRNA expression during early development. We performed an unbiased, genome-wide CRISPRi screen targeting thousands of lncRNA loci expressed during endoderm differentiation. While dozens of lncRNA loci were required for proper differentiation, most differentially expressed lncRNAs were not, supporting the necessity for functional screening instead of relying solely on gene expression analyses. In parallel, we developed a clustering approach to infer mechanisms of action of lncRNA hits based on a variety of genomic features. We subsequently identified and validated FOXD3-AS1 as a functional lncRNA essential for pluripotency and differentiation. Taken together, the cell lines and methodology described herein can be adapted to discover and characterize novel regulators of differentiation into any lineage.