Biology subjects

Buttitta, L.

Publications and source records attributed to Buttitta, L..

11 recordsLinked to original sources

bioRxiv2026

Mi-2 and E93 coordinate robust cell cycle exit with terminal differentiation through enhancer decommissioning

Many postmitotic tissues coordinate robust cell cycle exit with the progression of terminal differentiation. While the signals involved in initiating cell cycle exit during terminal differentiation for some tissues have been described, less is known about the mechanisms that maintain a stable, non-cycling state. We previously found that a subset of rate-limiting cell cycle genes are subject to developmentally controlled changes in chromatin accessibility after cell cycle exit, suggesting chromatin remodelers may play a key role in maintaining a robust postmitotic state during terminal differentiation. Here we show that the general chromatin remodeler Mi-2 is required to ensure a stable postmitotic state in Drosophila eyes and wings. Mi-2-mediated remodeling coordinates cell state transitions during development, as compromising Mi-2 function slows the chromatin accessibility and gene expression dynamics that are critical for tissue maturation. Mi-2 and a temporally expressed transcription factor involved in terminal differentiation, E93, each decommission chromatin at developmentally dynamic regions near a limited set of shared target genes, including the early differentiation factor Broad and the rate-limiting mitotic regulator string (cdc25c). Our evidence supports a model where Mi-2 and E93 act in vivo to decommission a newly identified distal string enhancer that is active during cell cycle exit, to coordinate the transition to a robust postmitotic state with the progression of terminal differentiation. Temporally regulated enhancer decommissioning at the string locus, depending on both an instructive temporal factor and an enzymatic remodeler, provides a molecular explanation for the long-term, nearly irreversible postmitotic state observed in many tissues as terminal differentiation progresses.

developmental biology↗

bioRxiv2025

Changes in cell size occur across brain regions in the aging Drosophila brain.

Aging is accompanied by complex cellular changes in the brain, yet alterations in cell size and brain volume during early adulthood remain poorly characterized. We used flow cytometry to examine cell size in the aging Drosophila melanogaster brain across genotypes, sexes, and brain regions, comparing the optic lobes (OL) and central brain (CB). We found substantial heterogeneity in brain cell sizes accompanied by a significant decrease in average cell sizes with age across genotypes, sexes, and regions. This reduction emerged early in adulthood and occurred in both the OL and CB, although the OL contained smaller cells than the CB. In parallel, we detected significant age-dependent accumulation of polyploid cells, defined as cells with a greater than 2C DNA content. Although polyploid cells were larger than diploid cells, they also exhibited an age-associated decline in average size. Adult-specific neuronal overexpression of Myc blunted the age-associated reduction in average cell size, indicating that this phenotype is modifiable by a known cell growth regulator. To determine whether cellular changes were accompanied by tissue-level alterations, we used micro-computed tomography to assess brain volume during early adulthood. In w1118 flies, brain volume decreased with age regardless of sex, consistent with the decrease we observe in average brain cell size. Together, these findings identify early brain-wide changes in cell size and volume during early adulthood in Drosophila melanogaster as prominent features of early brain aging and provide a foundation for investigating age-dependent cellular remodeling and its underlying mechanisms and functional consequences.

cell biology↗

bioRxiv2025

Evidence for Transcriptomic Conservation Between the Main Cells of the Drosophila Prostate-Like Accessory Gland and Basal Cells of the Mammalian Prostate.

The Drosophila accessory gland performs functions analogous to the mammalian prostate in production of seminal fluid components that are essential for male fertility. The mammalian prostate and Drosophila accessory glands share a similar tissue organization and structure. Both organs contain secretory epithelial cells forming a gland lumen, surrounded by a stroma with extracellular matrix enveloped by innervated muscle for organ contraction and fluid release. However, the Drosophila accessory gland secretory epithelium is postmitotic, polyploid and binucleate, and lacks a known stem cell population. By contrast, the mammalian prostate epithelium is made up of diploid luminal secretory cells and diploid basal cells that are maintained by at least two stem cell populations. Despite the differences in the tissues, it has been argued these tissues may share a deep homology based on the expression of conserved genes during development. Here we performed a cross-species comparative analysis using single-cell RNA sequencing data from adult tissues using data from the Drosophila Fly Cell Atlas and mammalian adult prostate single-cell datasets. Our analysis provides additional evidence of transcriptomic similarity between the main epithelial cells of the Drosophila prostate-like accessory gland and the basal epithelial cells of the mammalian prostate. While we do not know whether these similarities reflect shared evolutionary homology, or independently derived features due to shared tissue functions, our results strengthen the arguments that the Drosophila accessory gland can be used to effectively model aspects of human prostate biology and disease.

genomics↗

bioRxiv2025

A window of cell cycle plasticity enables imperfect regeneration of an adult postmitotic organ in Drosophila

The Drosophila ejaculatory duct (ED) is a secretory tissue of the male somatic reproductive system responsible for producing components of the seminal fluid which support fertility, serve antimicrobial functions and influence the physiological changes in the female after mating. The ED is a simple organ made up of secretory epithelial cells that are encased by extracellular matrix and a layer of innervated contractile muscle. The ED secretory epithelial cells are post-mitotic and lack known stem cells or progenitors in the adult, but they are not fully quiescent. They undergo a variant cell cycle called endoreplication immediately post-eclosion to increase organ size and protein synthesis capacity. Polyploid and post-mitotic tissues often face unique challenges in response to cell loss due to their inability to proliferate. Here, we show that the adult ED is capable of significant recovery after cell loss due to a combination of increased nuclear and cellular hypertrophy that partially restores tissue mass and organ function. The early cell cycle plasticity of this adult tissue is critical for this recovery, as older tissues that have few or no endocycles exhibit reduced capacity for recovery after cell loss. Together, our findings establish the Drosophila ED as a model to study post-mitotic polyploid tissue repair and highlight a combination of endocycles and hypertrophy as a key mechanism for functional regeneration in the absence of mitosis.

developmental biology↗

bioRxiv2024

Cell cycle re-entry in the aging Drosophila brain.

The brain is an organ comprised mostly of long-lived, quiescent cells that perform vital functions throughout an animals life. Due to the brains limited regenerative ability, these long-lived cells must engage unique mechanisms to cope with accumulated damage over time. We have shown that a subset of differentiated neuronal and glial cells in the fruit fly brain become polyploid during adulthood. Cell cycle re-entry in the brain has previously been associated with neurodegeneration, but there may be a more complex relationship between polyploidy and cell fitness in the brain. Here, we examine how known lifespan modifiers influence the accumulation of polyploidy in the aging fly brain. Flies aged at a low temperature, or with a low protein diet, accumulate polyploid cells in the brain more slowly than expected if this phenotype were solely regulated by lifespan mechanisms. Despite the slower accumulation of polyploid cells, animals under conditions that extend lifespan eventually reach similar levels of polyploidy in the brain as controls. Our work suggests known lifespan modifiers can influence the timing of cell cycle re-entry in the adult brain, indicating there is a flexible window of cell cycle plasticity in the aging brain.

developmental biology↗

bioRxiv2024

Post-eclosion growth in the Drosophila Ejaculatory Duct is driven by Juvenile Hormone signaling and is essential for male fertility

The Drosophila Ejaculatory duct (ED) is a secretory tissue of the somatic male reproductive system. The ED is involved in the secretion of seminal fluid components and ED-specific antimicrobial peptides that aid in fertility and the female post-mating response. The ED is composed of secretory epithelial cells surrounded by a layer of innervated contractile muscle. The ED grows in young adult males during the first 24h post-eclosion, but the cell cycle status of the ED secretory cells and the role of post-eclosion ED growth have been unexplored. Here, we show that secretory cells of the adult Drosophila ED undergo variant cell cycles lacking mitosis called the endocycle, that lead to an increase in the cell and organ size of the ED post eclosion. The cells largely exit the endocycle by day 3 of adulthood, when the growth of the ED ceases, resulting in a tissue containing cells of ploidies ranging from 8C-32C. The size of the ED directly correlates with the ploidy of the secretory cells, with additional ectopic endocycles increasing organ size. When endoreplication is compromised in ED secretory cells, it leads to reduced organ size, reduced protein synthesis and compromised fertility. We provide evidence that the growth and endocycling in the young adult male ED is dependent on Juvenile hormone (JH) signaling and we suggest that hormone-induced early adult endocycling is required for optimal fertility and function of the ED tissue. We propose to use the ED as a post-mitotic tissue model to study the role of polyploidy in regulating secretory tissue growth and function.

developmental biology↗

bioRxiv2022

A tissue dissociation method optimized for ATAC-seq and CUT&RUN in Drosophila pupal tissues.

Chromatin accessibility, histone modifications and transcription factor binding are highly dynamic during Drosophila metamorphosis and drive global changes in gene expression as larval tissues differentiate into adult structures. Unfortunately, the presence of pupa cuticle on many Drosophila tissues during metamorphosis prevents enzyme access to cells and has limited the use of enzymatic in situ methods for assessing chromatin accessibility and histone modifications. Here, we present a dissociation method for cuticle-bound pupal tissues that is optimized for use with ATAC-Seq and CUT&RUN to interrogate chromatin accessibility and histone modifications. We show this method provides comparable chromatin accessibility data to the non-enzymatic approach FAIRE-seq, with only a fraction of the amount of input tissue required. This approach is also compatible with CUT&RUN, which allows genome-wide mapping of histone modifications with less than 1/10th of the tissue input required for more conventional approaches such as Chromatin Immunoprecipitation Sequencing (ChIP-seq). Our protocol makes it possible to use newer, more sensitive enzymatic in situ approaches to interrogate gene regulatory networks during Drosophila metamorphosis.

developmental biology↗

bioRxiv2019

Polyploidy in the adult Drosophila brain

Long-lived cells such as terminally differentiated postmitotic neurons and glia must cope with the accumulation of damage over the course of an animals lifespan. How long-lived cells deal with ageing-related damage is poorly understood. Here we show that polyploid cells accumulate in the ageing adult fly brain and that polyploidy protects against DNA damage-induced cell death. Multiple types of neurons and glia that are diploid at eclosion, become polyploid with age in the adult Drosophila brain. The optic lobes exhibit the highest levels of polyploidy, associated with an elevated DNA damage response in this brain region with age. Inducing oxidative stress or exogenous DNA damage leads to an earlier onset of polyploidy, and polyploid cells in the adult brain are more resistant to DNA damage-induced cell death than diploid cells. Our results suggest polyploidy may serve a protective role for neurons and glia in ageing Drosophila melanogaster brains.

developmental biology↗

bioRxiv2019

Endocycling in the adult Drosophila accessory gland

The Drosophila melanogaster accessory gland is a functional analog of the mammalian prostate containing two secretory epithelial cell types, termed main and secondary cells. This tissue is responsible for making and secreting seminal fluid proteins and other molecules that contribute to successful reproduction. The cells of this tissue are bi-nucleate and polyploid, due to variant cell cycles that include endomitosis and endocycling during metamorphosis. Here we provide evidence of additional cell cycle variants in this tissue. We show that main cells of the gland are connected by ring canals that form after the penultimate mitosis and we describe an additional post-eclosion endocycle required for gland maturation that is dependent on juvenile hormone signaling. We present evidence that the main cells of the Drosophila melanogaster accessory gland undergo a unique cell cycle reprogramming throughout organ development that results in step-wise cell cycle truncations culminating in cells containing two octoploid nuclei with under-replicated heterochromatin in the mature gland. We propose this tissue as a model to study developmental and hormonal temporal control of cell cycle variants in terminally differentiating tissues.

developmental biology↗

bioRxiv2018

The coordination of terminal differentiation and cell cycle exit is mediated through the regulation of chromatin accessibility

During terminal differentiation most cells will exit the cell cycle and enter into a prolonged or permanent G0. Cell cycle exit is usually initiated through the repression of cell cycle gene expression by formation of a transcriptional repressor complex called DREAM. However when DREAM repressive function is compromised during terminal differentiation, additional unknown mechanisms act to stably repress cycling and ensure robust cell cycle exit. Here we provide evidence that developmentally programmed, temporal changes in chromatin accessibility at a subset of critical cell cycle genes acts to enforce cell cycle exit during terminal differentiation in the Drosophila melanogaster wing. We show that during terminal differentiation, chromatin closes at a set of pupal wing enhancers for the key rate-limiting cell cycle regulators cycE, e2f1 and stg. This closing coincides with wing cells entering a robust postmitotic state that is strongly refractory to cell cycle re-activation. When cell cycle exit is genetically disrupted, chromatin accessibility at cell cycle genes remains largely unaffected and the closing of enhancers at cycE, e2f1 and stg proceeds independent of the cell cycling status. Instead, disruption of cell cycle exit leads to changes in accessibility and expression of a subset of hormone-induced transcription factors involved in the progression of terminal differentiation. Our results uncover a mechanism that acts as a cell cycle-independent timer to limit aberrant cycling in terminally differentiating tissues. In addition, we provide a new molecular description of the cross-talk between cell cycle exit and terminal differentiation during metamorphosis.

developmental biology↗

bioRxiv2017

Chromatin organization changes during the establishment and maintenance of the postmitotic state

BackgroundGenome organization changes during development as cells differentiate. Chromatin motion becomes increasingly constrained and heterochromatin clusters as cells become restricted in their developmental potential. These changes coincide with slowing of the cell cycle, which can also influence chromatin organization and dynamics. Terminal differentiation is often coupled with permanent exit from the cell cycle and existing data suggests a close relationship between a repressive chromatin structure and silencing of the cell cycle in postmitotic cells. Here we examine the relationship between chromatin organization, terminal differentiation and cell cycle exit.\n\nResultsWe focused our studies on the Drosophila wing, where epithelial cells transition from active proliferation to a postmitotic state in a temporally controlled manner. We find there are two stages of G0 in this tissue, a flexible G0 period where cells can be induced to re-enter the cell cycle under specific genetic manipulations and a state we call \"robust\", where cells become strongly refractory to cell cycle re-entry. Compromising the flexible G0 by driving ectopic expression of cell cycle activators causes a global disruption of the clustering of heterochromatin-associated histone modifications such as H3K27 trimethylation and H3K9 trimethylation, as well as their associated repressors, Polycomb and heterochromatin protein 1(HP1). However, this disruption is reversible. When cells enter a robust G0 state, even in the presence of ectopic cell cycle activity, clustering of heterochromatin associated modifications are restored. If cell cycle exit is bypassed, cells in the wing continue to terminally differentiate, but heterochromatin clustering is severely disrupted. Heterochromatin-dependent gene silencing does not appear to be required for cell cycle exit, as compromising the H3K27 methyltransferase Enhancer of zeste, and/or HP1 cannot prevent the robust cell cycle exit, even in the face of normally oncogenic cell cycle activities.\n\nConclusionsHeterochromatin clustering during terminal differentiation is a consequence of cell cycle exit, rather than differentiation. Compromising heterochromatin-dependent gene silencing does not disrupt cell cycle exit.

developmental biology↗