Biology subjects

Weake, V. M.

Publications and source records attributed to Weake, V. M..

10 recordsLinked to original sources

bioRxiv2026

Defining cell type-specific diurnal gene expression in the mouse retina

The mammalian retina exhibits daily oscillations in gene expression that contribute to retinal physiology and visual function. However, the extent to which these temporal patterns differ among retinal cell types remains incompletely understood. We performed single-nucleus RNA sequencing in the mouse retina over the day and night, identifying rhythmic genes in the six major retinal cell types. Distinct diurnal gene expression patterns across photoreceptors, interneurons, retinal ganglion cells, and Muller glia correspond to their diverse functions across the light cycle. Even core circadian clock genes exhibited differences in paralog expression and rhythmicity across the retina. In contrast, dopamine signaling via the DRD4 receptor is highly rhythmic across most retinal cell types, suggesting dopamine signaling may be a key rhythmicity cue in the retina under diurnal conditions. This dataset highlights the complexity of daily transcriptional regulation in the retina and provides a resource to explore cell-type-specific diurnal rhythms.

neuroscience↗

bioRxiv2026

Chromatin states coordinate diurnal gene expression in the Drosophila retina

Daily biological rhythms enable organisms to anticipate and respond to predictable diurnal environmental changes. While the circadian clock maintains rhythms under constant conditions, it remains unclear what gates the time-of-day expression of genes that are rhythmic under light:dark cycles. In the retina, restricting biological processes to specific times of the day is crucial for homeostasis because disrupting these rhythms causes retinal degeneration. We show that in light-sensitive photoreceptor neurons of Drosophila, the circadian transcription factors only account for the timing of a minority of rhythmic genes. Instead, integrative analysis of the daily chromatin landscape and transcription factor footprints reveals that promoter, enhancer, or repressive signatures strongly correspond with phase. Further, genetic modulation of the repressive and promoter chromatin states altered gene expression in a phase-specific manner. Together, our data support a model where the genome is organized into discrete chromatin signatures that collaborate with transcription factors to restrict diurnal transcription to distinct phases of the day.

genomics↗

bioRxiv2025

PBRM1-Dependent PBAF Targeting is Required for EMT and Metastasis in Breast Cancer

SWI/SNF chromatin remodelers utilize ATP to mobilize nucleosomes on DNA and are represented by three biochemically distinct subcomplexes, the more abundant cBAF and the less abundant PBAF and GBAF subcomplexes. Patient mutations and genetic studies have identified important roles for PBAF subunits in development and disease; however, relating PBAF-mediated phenotypes to biochemical function in chromatin regulation and gene expression has been challenging. Further complicating matters, cell-based systems often do not reflect the phenotypes and genotypes observed with PBAF mutations in vivo. Here we show that the PBRM1 subunit of PBAF is critical for the completion of TGFB1-mediated epithelial-mesenchymal transition of mammary cells in vitro as well as the metastasis of murine breast cancers in vivo. Using epigenomics to profile different stages of EMT, we find that PBRM1 is necessary for targeting PBAF to inducible promoters marked by H3K14ac alone. We further find that PBRM1 facilitates DNA accessibility at sites bound by TGF{beta}1-inducible transcription factors, such as Atf3, for the induction of genes involved in migration, cell survival, and inflammation. Our model allows us to separate constitutive vs inducible gene expression to help explain some of the context-dependent phenotypes observed with PBRM1 deletion. In addition, we provide evidence that while PBRM1 deletions can promote the initiation of certain cancers in early stages, PBAF may be a vulnerability in late-stage metastatic cancers.

molecular biology↗

bioRxiv2025

Broad epigenetic shifts in the aging Drosophila retina contribute to its altered rhythmic transcriptome.

Alterations in biological rhythms are a common feature of aging, and disruption of circadian rhythms can exacerbate age-associated pathologies. The retina is critical for detecting light for both vision and for transmitting time-of-day information to the brain, synchronizing rhythms throughout the body. Disruption of circadian rhythms by manipulating the molecular clock leads to premature retinal degeneration in flies and mice, and gene expression rhythms are disrupted in models of age-associated ocular disease. Despite this, it is unknown how or why the gene expression rhythms of the retina change with age. Here, we show that [~]70% of the Drosophila transcriptome is rhythmically expressed throughout the diurnal cycle, with [~]40% of genes showing altered rhythms with age. These transcriptome-wide changes in aging photoreceptors are accompanied by shifts in the rhythmic patterns of RNA Polymerase II (Pol II) occupancy, histone H3 lysine 4 (H3K4) methylation, and chromatin accessibility, without major changes in occupancy of the circadian clock transcription factors Clock (Clk) and Cycle (Cyc). Instead, aging decreases genome-wide levels of several different histone methyl marks including H3K4 methylation, whose relative levels across the day correlate with the phase of rhythmic gene expression. Moreover, individual knockdown of the three H3K4 methyltransferases in young photoreceptors results in massive disruptions to rhythmic gene expression that resemble those observed during aging. We conclude that there are broad epigenetic shifts in the aging retina, including decreased histone methylation, that contribute to changes in biological rhythms even in the presence of a robust molecular circadian clock.

molecular biology↗

bioRxiv2025

One-carbon metabolism enzyme Ahcy is a redox sensor that modulates gene expression to protect against light stress-induced retinal degeneration

One-carbon metabolism influences gene expression by providing methyl units for DNA, RNA, and histone methylation. Robust methylation requires rapid hydrolysis of the methylation by-product S-adenosylhomocysteine (SAH) by S-adenosylhomocysteinase (Ahcy). Here, we show Ahcy is a redox-sensitive enzyme that is inhibited by oxidation of a conserved cysteine, C195, in vitro and in vivo. Transient oxidation of Ahcy is neuroprotective in a Drosophila light stress model where it results in rapid gene expression changes and protects against retinal degeneration. Thus, redox sensing by the one-carbon metabolic enzyme Ahcy enables rapid changes in gene expression in response to changes in redox homeostasis.

biochemistry↗

bioRxiv2022

Establishing the contribution of active histone methylation marks to the aging transcriptional landscape of Drosophila photoreceptors

Studies in multiple organisms have shown that aging is accompanied by several molecular phenotypes that include dysregulation of chromatin. Since chromatin regulates DNA-based processes such as transcription, alterations in chromatin modifications could impact the transcriptome and function of aging cells. In flies, as in mammals, the aging eye undergoes changes in gene expression that correlate with declining visual function and increased risk of retinal degeneration. However, the causes of these transcriptome changes are poorly understood. Here, we profiled chromatin marks associated with active transcription in the aging Drosophila eye to understand how chromatin modulates transcriptional outputs. We found that both H3K4me3 and H3K36me3 globally decrease across all actively expressed genes with age. However, we found no correlation with changes in differential gene expression. Downregulation of the H3K36me3 methyltransferase Set2 in young photoreceptors revealed significant changes in splicing events that overlapped significantly with those observed in aging photoreceptors. These overlapping splicing events impacted multiple genes involved in phototransduction and neuronal function. Since proper splicing is essential for visual behavior, and because aging Drosophila undergo a decrease in visual function, our data suggest that H3K36me3 plays a role in maintaining visual function in the aging eye through regulating alternative splicing.

genomics↗

bioRxiv2021

The Clock:cycle complex is a major transcriptional regulator of Drosophila photoreceptors that protects the eye from retinal degeneration and oxidative stress

The aging eye experiences physiological changes that include decreased visual function and increased risk of retinal degeneration. Although there are transcriptomic signatures in the aging retina that correlate with these physiological changes, the gene regulatory mechanisms that contribute to cellular homeostasis during aging remain to be determined. Here, we integrated ATAC-seq and RNA-seq data to identify 61 transcription factors that showed differential activity in aging Drosophila photoreceptors. These 61 age-regulated transcription factors include two circadian regulators, Clock and cycle, that showed sustained increases in activity during aging. When we disrupted Clock activity in adult photoreceptors, we observed changes in expression of 15 - 20% of genes including key components of the phototransduction machinery and many eye-specific transcription factors. Using ATAC-seq, we showed that loss of Clock activity leads to changes in activity of 31 transcription factors and causes a progressive decrease in global levels of chromatin accessibility in photoreceptors. Supporting a key role for Clock-dependent transcription in the eye, disruption of Clock activity in photoreceptors also induced light-dependent retinal degeneration and increased oxidative stress, independent of light exposure. Together, our data suggests that the circadian regulators Clock and cycle act as neuroprotective factors in the aging eye by directing gene regulatory networks that maintain expression of the phototransduction machinery and counteract oxidative stress.

genomics↗

bioRxiv2021

Proper control of R-loop homeostasis is required for maintenance of gene expression and neuronal function during aging

Age-related loss of cellular function and increased cell death are characteristic hallmarks of aging. While defects in gene expression and RNA metabolism have been linked with age-associated human neuropathies, it is not clear how the changes that occur in aging neurons contribute to loss of gene expression homeostasis. R-loops are RNA-DNA hybrids that typically form co-transcriptionally via annealing of the nascent RNA to the template DNA strand, displacing the non-template DNA strand. Dysregulation of R-loop homeostasis has been associated with both transcriptional impairment and genome instability. Importantly, a growing body of evidence links R-loop accumulation with cellular dysfunction, increased cell death and chronic disease onset. Here, we characterized the R-loop landscape in aging Drosophila melanogaster photoreceptor neurons and showed that bulk R-loop levels increased with age. Further, genome-wide mapping of R-loops revealed that transcribed genes accumulated R-loops over gene bodies during aging, which correlated with decreased expression of long and highly expressed genes. Importantly, while photoreceptor-specific down-regulation of Top3{beta}, a DNA/RNA topoisomerase associated with R-loop resolution, lead to decreased visual function, overexpression of Top3{beta} or nuclear-localized RNase H1, which resolves R-loops, enhanced positive light response during aging. Together, our studies highlight the functional link between dysregulation of R-loop homeostasis, gene expression and visual function during aging.

molecular biology↗

bioRxiv2021

Quantitative proteomic and metabolomic profiling reveals altered mitochondrial metabolism and folate biosynthesis pathways in the aging Drosophila eye

Aging is associated with increased risk of ocular disease, suggesting that age-associated molecular changes in the eye increase its vulnerability to damage. Although there are common pathways involved in aging at an organismal level, different tissues and cell types exhibit specific changes in gene expression with advanced age. Drosophila melanogaster is an established model system for studying aging and neurodegenerative disease, that also provides a valuable model for studying age-associated ocular disease. Flies, like humans, exhibit decreased visual function and increased risk of retinal degeneration with age. Here, we profiled the aging proteome and metabolome of the Drosophila eye, and compared these data with age-associated transcriptomic changes from both eyes and photoreceptors to identify alterations in pathways that could lead to age-related phenotypes in the eye. Notably, the proteomic and metabolomic changes observed in the aging eye are distinct from those observed in the head or whole fly, suggesting that tissue-specific changes in protein abundance and metabolism occur in the aging fly. Our integration of the proteomic, metabolomic and transcriptomic data reveals that changes in metabolism, potentially due to decreases in availability of B vitamins, together with chronic activation of the immune response, may underpin many of the events observed in the aging Drosophila eye. We propose that targeting these pathways in the genetically tractable Drosophila system may help to identify potential neuroprotective approaches for neurodegenerative and age-related ocular diseases.

biochemistry↗

bioRxiv2021

In vivo tissue-specific chromatin profiling in Drosophila melanogaster using GFP-tagged nuclei

AbstractThe chromatin landscape defines cellular identity in multicellular organisms with unique patterns of DNA accessibility and histone marks decorating the genome of each cell type. Thus, profiling the chromatin state of different cell types in an intact organism under disease or physiological conditions can provide insight into how chromatin regulates cell homeostasis in vivo. To overcome the many challenges associated with characterizing chromatin state in specific cell types, we developed an improved approach to isolate Drosophila nuclei tagged with GFP expressed under Gal4/UAS control. Using this protocol, we profiled chromatin accessibility using Omni-ATAC, and examined the distribution of histone marks using ChIP-seq and CUT&Tag in adult photoreceptor neurons. We show that the chromatin landscape of photoreceptors reflects the transcriptional state of these cells, demonstrating the quality and reproducibility of our approach for profiling the transcriptome and epigenome of specific cell types in Drosophila.

genomics↗