Biology subjects

Greenberg, A.

Publications and source records attributed to Greenberg, A..

8 recordsLinked to original sources

bioRxiv2026

Constitutive ATR Signaling Enforces Domain-Wide Origin Synchrony

Eukaryotic DNA replication relies on organized spatial-temporal domains, yet how neighboring origins are coordinated at the single-molecule level remains unresolved. Combining single-molecule DNA combing with super-resolution microscopy, we demonstrate that during unperturbed early S-phase, adjacent replication origins across diverse vertebrate species fire synchronously as a coordinated unit. This domain-wide temporal harmony is continuously enforced by a baseline, constitutive ATR CHK1 signaling axis that restrains local CDK2 and CDC7 kinase activities. Strikingly, replication stress dismantles this coordination, shifting origin activation toward an asynchronous, replicon-autonomous mode. Mechanistically, stress-induced phosphorylation of the cohesin subunit SMC1A promotes an SMC1A ATR complex that sequesters ATR at stalled replication forks. This localized sequestration creates a regional ATR loss-of-function phenotype, relieving CDK2/CDC7 suppression and driving premature, uncoordinated origin firing. Super-resolution imaging shows that ATR sequestration structurally reorganizes replication foci, converting coordinated clustered foci into fragmented, lower-density clusters. Together, our findings uncover a baseline role for constitutive ATR signaling in enforcing domain synchrony and reveal a molecular switch that dynamically decouples origin activation during genomic stress to safeguard chromosome duplication.

molecular biology↗

bioRxiv2026

A ML-framework for the discovery of next-generation IBD targets using a harmonized single-cell atlas of patient tissue

Target discovery for IBD has traditionally relied on genetic associations, which lack the cellular resolution needed to identify novel, actionable, cell type-specific disease pathways. Here, we describe an integrated analytical and experimental framework that leverages harmonized single-cell data to systematically discover novel therapeutic strategies for IBD. We used AMICA DBTM, Immunais harmonized database of single-cell RNA datasets to construct a harmonized 1 million single-cell atlas of the human intestine. We applied a machine learning framework (Immune Patient Representation, IPR) to identify disease-associated transcriptional programs and cell type-specific gene targets. Candidate targets were prioritized using atlas-derived metrics, refined using custom criteria emphasizing translational actionability, and validated across independent clinical cohorts. Select candidates were evaluated in human primary-cell models reflecting the targets cell-type context. The IPR framework identified 85 disease-associated transcriptional programs and ranked 400 cell type-specific target genes across immune and stromal lineages. Disease-associated programs were interpreted using a structured AI-assisted reasoning framework for structured biological reasoning, linking them to IBD-relevant pathways and guiding the identification of novel, promising gene targets. Functional validation of two cell-type-specific candidates, PTGIR in myeloid cells and IL6ST in fibroblasts, confirmed the reduction of inflammatory and fibrotic pathways linked to IBD pathology. Multi-omic profiling and projection of in vitro phenotypes to patient datasets demonstrated the reversal of disease-associated programs via mechanisms distinct from those of existing biologics. Our single-cell anchored, machine-learning framework integrates in silico discovery with experimental validation, revealing new cell type-specific therapeutic opportunities and supporting a scalable approach for precision target discovery in IBD and other immune-mediated diseases.

bioinformatics↗

bioRxiv2025

The landscape of allele-specific expression in human kidneys

Allele-specific expression (ASE), the preferential expression of one gene copy, is a key mechanism of genomic regulation. However, its role in human kidney disease remains poorly understood. In this study, we generated a high-quality, genome-wide ASE map using paired whole-genome sequencing and RNA-seq from microdissected glomerular (GLOM) and tubulointerstitial (TUBE) compartments of patients with proteinuric kidney disease. We showed that the majority of common ASE events were deterministic and sequence-mediated. We also found that diseased kidneys exhibited significantly more ASE in GLOM than TUBE, compared to controls. Unexpectedly, higher ASE in GLOM than TUBE was significantly associated with improved kidney disease outcomes in the disease cohort. Differential gene expression analysis suggested this was the result of an active, protective transcriptional response, including ribosome and ATP synthesis upregulation, rather than pathogenic dysregulation. Our work reveals glomerular ASE as a marker of adaptive transcriptional activity in proteinuric kidney disease.

genomics↗

bioRxiv2023

Dual impact of PTEN mutation on CSF dynamics and cortical networks via the dysregulation of neural precursors and their interneuron descendants

Expansion of the cerebrospinal fluid (CSF)-filled cerebral ventricles (ventriculomegaly) is the quintessential feature of congenital hydrocephalus (CH) but also seen in autism spectrum disorder (ASD) and several neuropsychiatric diseases. PTEN is frequently mutated in ASD; here, we show PTEN is a bona fide risk gene for the development of ventriculomegaly, including neurosurgically-treated CH. Pten-mutant hydrocephalus is associated with aqueductal stenosis due to the hyperproliferation of periventricular Nkx2.1+ neural precursors (NPCs) and CSF hypersecretion from inflammation-dependent choroid plexus hyperplasia. The hydrocephalic Pten-mutant cortex exhibits ASD-like network dysfunction due to impaired activity of Nkx2.1+ NPC-derived inhibitory interneurons. Raptor deletion or post-natal Everolimus corrects ventriculomegaly, rescues cortical deficits, and increases survival by antagonizing mTORC1-dependent Nkx2.1+ cell pathology. These results implicate a dual impact of PTEN mutation on CSF dynamics and cortical networks via the dysregulation of NPCs and their interneuron descendants. These data identify a non-surgical treatment target for hydrocephalus and have implications for other developmental brain disorders. HIGHLIGHTSO_LIPTEN de novo mutations are associated with cerebral ventriculomegaly in autism spectrum disorder (ASD) and congenital hydrocephalus (CH). C_LIO_LIPten-mutant hydrocephalus is associated with aqueductal stenosis due to the hyperproliferation of medial ganglionic eminence Nkx2.1+ neural precursors and CSF hypersecretion from inflammation-induced choroid plexus hyperplasia. C_LIO_LIThe hydrocephalic Pten-mutant cortex exhibits ASD-like network dysfunction due to impaired activity of Nkx2.1+ NPC-derived inhibitory interneurons. C_LIO_LImTORC1 inhibition via Raptor deletion or early post-natal treatment with rapamycin or everolimus increases survival and ameliorates Pten-mutant ventriculomegaly and cortical pathology. C_LI

neuroscience↗

bioRxiv2022

Mapping genomic regulation of kidney disease and traits through high-resolution and interpretable eQTLs

Expression quantitative trait locus (eQTL) studies illuminate genomic variants that regulate specific genes and contribute to fine-mapped loci discovered via genome-wide association studies (GWAS). Efforts to maximize their accuracy are ongoing. Using 240 glomerular (GLOM) and 311 tubulointerstitial (TUBE) micro-dissected samples from human kidney biopsies, we discovered 5,371 GLOM and 9,787 TUBE eQTLs by incorporating kidney single-nucleus open chromatin data and transcription start site distance as an "integrative prior" for Bayesian statistical fine mapping. The use of an integrative prior resulted in higher resolution eQTLs illustrated by (1) smaller numbers of variants in credible sets with greater confidence, (2) increased enrichment of partitioned heritability for GWAS of two kidney traits, (3) an increased number of variants colocalized with the GWAS loci, and (4) enrichment of computationally predicted functional regulatory variants. A subset of variants and genes were validated experimentally in vitro and using a Drosophila nephrocyte model. More broadly, this study demonstrates that tissue-specific eQTL maps informed by single-nucleus open chromatin data have enhanced utility for diverse downstream analyses.

genomics↗

bioRxiv2021

Cancer mutational processes vary in their association with replication timing and chromatin accessibility

BackgroundCancer somatic mutations are the product of multiple mutational and repair processes, which are tightly associated with DNA replication. Distinctive patterns of somatic mutations accumulation in tumors, termed mutational signatures, are indicative of processes the tumors underwent. While tumor mutational load is correlated with late replicating regions and spatial genome organization, much is unknown about the association of many different mutational processes and replication timing, and the interplay with chromatin structure remains an open question. MethodsWe systematically analyzed the mutational landscape of 2,787 WGS tumors from 32 different tumor types separately for early and late replicating regions. We used sequence context normalization and chromatin data to account for sequence and chromatin accessibility differences between early and late replicating regions. Moreover, we expanded the signature analyses to doublet base substitutions and small insertions and deletions by developing an artificial genomes-based approach to account for sequence differences between various genomic regions. ResultsWe revealed the replication timing (RT) association of single base, doublet base and small insertions and deletions mutational signatures. The association is signature specific: some are associated with early or late replication (such as UV-exposure signatures SBS7b and SBS7a, respectively) and others have no association. Most associations exist even after normalizing for genome accessibility. We further developed a focused mutational signature identification approach, which uses RT information to improve signature identification, and found that SBS16, which is biased towards early replication, is strongly associated with better survival rates in liver cancer. ConclusionsOur comprehensive analyses enabled a more robust classification of RT association of single base, doublet base and indels signatures. By doing so, we demonstrated a variation in the association with RT, as many mutational processes biased towards either early or late replication timing, and others have an equal RT distribution. These associations were independent from chromatin accessibility in most cases. This work highlights that restricting signatures analyses to concise genomic regions improves identification of signatures, such as SBS16, and demonstrates its clinically relevance as a predictor of improved survival of liver cancer patients.

genomics↗

bioRxiv2020

Perilipin2 down-regulation in beta cells impairs insulin secretion under nutritional stress and damages mitochondria

Perilipin 2 (PLIN2) is the lipid droplet (LD) protein in {beta} cells that increases under nutritional stress. Down-regulation of PLIN2 is often sufficient to reduce LD accumulation. To determine whether PLIN2 positively or negatively affects {beta} cell function under nutritional stress, PLIN2 was down-regulated in mouse {beta} cells, INS1 cells, and human islet cells. {beta} cell specific deletion of PLIN2 in mice on a high fat diet reduced glucose-stimulated insulin secretion (GSIS) in vivo and in vitro. Down-regulation of PLIN2 in INS1 cells blunted GSIS after 24 h incubation with 0.2 mM palmitic acids. Down-regulation of PLIN2 in human pseudoislets cultured at 5.6 mM glucose impaired both phases of GSIS, indicating that PLIN2 is critical for GSIS. Down-regulation of PLIN2 decreased specific OXPHOS proteins in all three models and reduced oxygen consumption rates in INS1 cells and mouse islets. Moreover, we found that PLIN2 deficient INS1 cells increased the distribution of a fluorescent oleic acid analog to mitochondria and showed signs of mitochondrial stress as indicated by susceptibility to fragmentation and alterations of acyl-carnitines and glucose metabolites. Collectively, PLIN2 in {beta} cells have an important role in preserving insulin secretion, {beta} cell metabolism and mitochondrial function under nutritional stress.

molecular biology↗

bioRxiv2017

New waves: Rhythmic electrical field stimulation systematically alters spontaneous slow dynamics across mouse neocortex

The signature rhythm of slow-wave forebrain activity is the large amplitude, slow oscillation (SO: ~1 Hz) made up of alternating synchronous periods of depolarizing and hyperpolarizing states at the single cell and network levels. On each wave, the SO originates at a unique location and propagates across the neocortex. Attempts to manipulate SO activity using electrical fields have been shown to entrain cortical networks and enhance memory performance. However, neural activity during this manipulation has remained elusive due to methodological issues in typical electrical recordings. Here we use voltage-sensitive dye (VSD) imaging in a bilateral cortical preparation of urethane-anesthetized mice to track SO cortical activity and its modulation by sinusoidal electrical field stimulation applied to frontal regions. We show that under spontaneous conditions, the SO propagates in two main opposing directional patterns along an anterior lateral - posterior medial axis. Rhythmic field stimulation alters spontaneous propagation to reflect activity that repeats cycle after cycle with distributed and varied anterior initiation zones and a consistent termination zone in the posterior somatosensory cortex. Our results show that slow electrical field stimulation stereotypes ongoing slow cortical dynamics during sleep-like states.\n\nAuthor ContributionsA.G., J.K.A., M.H.M. and C.T.D. designed the study. A.G. and J.K.A. performed the experiments. A.G. analyzed the data. A.G. wrote the manuscript, which all authors commented on and edited. C.T.D. and M.H.M. supervised the study.

neuroscience↗