Biology subjects

Tootle, T. L.

Publications and source records attributed to Tootle, T. L..

11 recordsLinked to original sources

bioRxiv2026

Prostaglandins regulate the nucleoskeleton during Drosophila border cell migration

The nucleoskeleton, which is comprised of Lamin A (stiffer), Lamin B, and Lamin interacting proteins, including Emerin, controls nuclear stiffness. Nuclear stiffness regulates 3D single cell migration, but its roles in collective cell migration remain unclear. To define the roles of the nucleoskeleton during collective migration we use Drosophila border cell migration. During migration the nucleoskeleton remodels. Throughout migration, Lamin A is predominantly in the nucleoskeleton of the polar cells, whereas Emerin is progressively reduced in the nucleoskeletons of both the border and polar cells, and Lamin B increases in the border cell nucleoskeleton. Further, the border cell nucleoskeleton is polarized; Lamin B is enriched in the front of the cluster while Emerin is enriched in the back. These nucleoskeletal changes require prostaglandin (PG) signaling. When PG signaling is lost, border cell migration is delayed, Lamin A and Emerin are prevalent within the border cell nucleoskeletons throughout migration and nucleoskeletal polarity is lost. Further, overexpression of Lamin A R237P in the border cells delays migration. These data reveal that border cell cluster nucleoskeletal remodeling requires PG signaling and support that this remodeling facilitates invasive, collective migration. Similar PG regulation of the nucleoskeleton likely promotes collective migration across organisms and contexts. Significance StatementO_LINucleoskeletal remodeling is critical for 3D single cell migration, but its roles in collective migration are poorly understood. C_LIO_LIDuring Drosophila border cell migration, the nucleoskeleton remodels and exhibits polarity that suggests the nuclei are softer in the front and stiffer in the back of the cluster. PG signaling is required for these nucleoskeletal changes and on-time border cell migration. Overexpression of Lamin A R237P in the border cells impairs migration. C_LIO_LIThese results demonstrate for the first time that nucleoskeletal remodeling occurs during an in vivo, collective cell migration and identify PG signaling as a novel regulator of the nucleoskeleton. C_LI

cell biology↗

bioRxiv2026

ATGL-mediated lipid droplet lipolysis promotes collective migration in Drosophila

While lipid droplets (LDs), dynamic organelles central to lipid and energy homeostasis, are implicated in cancer cell migration, their roles during collective cell migration remain unknown. We use Drosophila border cell migration as an in vivo model of invasive, collective cell migration to dissect the roles of LDs and the conserved LD lipase, Adipose Triglyceride Lipase (ATGL). Border cell LDs undergo dynamic changes and decrease in volume by the end of migration. Loss of ATGL increases LD volume, whereas border cell overexpression depletes LDs. Loss, border cell knockdown or overexpression of ATGL delays migration and blocks delamination. Further, loss of ATGL disrupts border cell mitochondria - it alters morphology, reduces membrane potential and increases reactive oxygen species. These results demonstrate that tight regulation of lipid mobilization from LDs, including for energy production, drives delamination and collective migration. Our findings not only have the potential to inform how cancer cells exploit LDs to promote their invasive behaviors but also highlight the crucial role of LDs in migration during development, hinting at their broader significance in diverse migratory contexts.

cell biology↗

bioRxiv2025

The lipid droplet protein Jabba promotes actin remodeling downstream of prostaglandin signaling during Drosophila oogenesis

Growing evidence supports that lipid droplets (LDs) are critical for producing high-quality oocytes. However, the functions of LDs during oocyte development remain largely unknown. Using Drosophila oogenesis as a model, we previously discovered the LD-associated Adipose Triglyceride Lipase (ATGL) promotes actin remodeling necessary for oocyte development by providing the substrate for producing lipid signals termed prostaglandins (PGs). Here we find that Jabba, a LD-associated protein best known for its role in anchoring other proteins to LDs, also promotes PG-dependent actin remodeling. Overexpression of Jabba results in thickened cortical actin and excessive actin bundles, whereas loss of Jabba results in cortical actin breakdown and severely defective actin bundle formation. We find that Jabba regulates actin remodeling independently of ATGL but in conjunction with PG signaling. These data support that there are two PG signaling pathways that promote actin remodeling: one PG pathway that is dependent on ATGL and the other requires Jabba. Overexpression of Jabba rescues the actin defects when PG signaling is lost. Together these data lead to the model that PGs produced independently of ATGL positively regulate Jabba to promote actin remodeling necessary for follicle morphogenesis and the production of a fertilization competent oocyte. Significance statementO_LIAcross organisms, lipid droplets accumulate during oocyte development and are implicated in fertility. The functions of lipid droplets during oogenesis are poorly understood. C_LIO_LIThe authors use the genetic tools and well-characterized process of Drosophila oogenesis to reveal that Jabba, a lipid droplet anchoring protein, is a new downstream effector of prostaglandin signaling and promotes actin remodeling necessary for producing a fertilization competent oocyte. C_LIO_LIThe results extend prior studies connecting lipid droplet proteins, prostaglandins, and actin remodeling, providing insight into how these critical conserved factors contribute to high-quality oocytes. C_LI

cell biology↗

bioRxiv2024

Lipid droplet protein Perilipin 2 is critical for the regulation of insulin secretion through beta cell lipophagy and glucagon expression in pancreatic islets

Knockdown (KD) of lipid droplet (LD) protein perilipin 2 (PLIN2) in beta cells impairs glucose-stimulated insulin secretion (GSIS) and mitochondrial function. Here, we addressed a pathway responsible for compromised mitochondrial integrity in PLIN2 KD beta cells. In PLIN2 KD human islets, mitochondria were fragmented in beta cells but not in alpha cells. Glucagon but not insulin level was elevated. While the formation of early LDs followed by fluorescent fatty acids (FA) analog Bodipy C12 (C12) was preserved, C12 accumulated in mitochondria over time in PLIN2 KD INS-1 cells. A lysosomal acid lipase inhibitor Lali2 prevented C12 transfer to mitochondria, mitochondrial fragmentation, and the impairment of GSIS. Direct interactions between LD-lysosome and lysosome-mitochondria were increased in PLIN2 KD INS-1 cells. Thus, FA released from LDs by microlipophagy cause mitochondrial changes and impair GSIS in PLIN2 KD beta cells. Interestingly, glucolipotoxic condition (GLT) caused C12 accumulation and mitochondrial fragmentation similar to PLIN2 KD in beta cells. Moreover, Lali2 reversed mitochondrial fragmentation and improved GSIS in human islets under GLT. In summary, PLIN2 regulates microlipophagy to prevent excess FA flux to mitochondria in beta cells. This pathway also contributes to GSIS impairment when LD pool expands under nutrient load in beta cells.

molecular biology↗

bioRxiv2024

Nuclear actin is a critical regulator of Drosophila female germline stem cell maintenance

Nuclear actin has been implicated in regulating cell fate, differentiation, and cellular reprogramming. However, its roles in development and tissue homeostasis remain largely unknown. Here we uncover the role of nuclear actin in regulating stemness using Drosophila ovarian germline stem cells (GSCs) as a model. We find that the localization and structure of nuclear actin is dynamic in the early germ cells. Nuclear actin recognized by anti-actin C4 is found in both the nucleoplasm and nucleolus of GSCs. The polymeric nucleoplasmic C4 pool is lost after the 2-cell stage, whereas the monomeric nucleolar pool persists to the 8-cell stage, suggesting that polymeric nuclear actin may contribute to stemness. To test this idea, we overexpressed nuclear targeted actin constructs to alter nuclear actin polymerization states in the GSCs and early germ cells. Increasing monomeric nuclear actin, but not polymerizable nuclear actin, causes GSC loss that ultimately results in germline loss. This GSC loss is rescued by simultaneous overexpression of monomeric and polymerizable nuclear actin. Together these data reveal that GSC maintenance requires polymeric nuclear actin. This polymeric nuclear actin likely plays numerous roles in the GSCs, as increasing monomeric nuclear actin disrupts nuclear architecture causing nucleolar hypertrophy, distortion of the nuclear lamina, and heterochromatin reorganization; all factors critical for GSC maintenance and function. These data provide the first evidence that nuclear actin, and in particular, its ability to polymerize, are critical for stem cell function and tissue homeostasis in vivo.

developmental biology↗

bioRxiv2021

Lipid droplets regulate actin remodeling via prostaglandin signaling during Drosophila oogenesis

A key factor controlling oocyte quality and fertility is lipids. Even though lipid droplets (LDs) are crucial regulators of lipid metabolism, their roles in fertility are poorly understood. During Drosophila oogenesis, LD accumulation in nurse cells coincides with dynamic actin remodeling necessary for late-stage follicle morphogenesis and fertility. Loss of the LD-associated Adipose Triglyceride Lipase (ATGL) disrupts both actin bundle formation and cortical actin integrity, an unusual phenotype also seen when Pxt, the enzyme responsible for prostaglandin (PG) synthesis, is missing. Dominant genetic interactions and PG treatment of follicles in vitro reveal that ATGL and Pxt act in the same pathway to regulate actin remodeling, with ATGL upstream of Pxt. Further, lipidomic analysis detects arachidonic acid (AA) containing triglycerides in ovaries. Because AA is the substrate for Pxt, we propose that ATGL releases AA from LDs to drive PG synthesis necessary for follicle development. We also find that exogenous AA is toxic to follicles in vitro, and LDs modulate this toxicity. This leads to the model that LDs both sequester AA to limit toxicity, and release AA via ATGL to drive PG production. We speculate that the same pathways are conserved across organisms to regulate oocyte development and promote fertility.

cell biology↗

bioRxiv2021

Fascin limits Myosin activity within Drosophila border cells to control substrate stiffness and promote migration

A key regulator of collective cell migrations, which drive development and cancer metastasis, is substrate stiffness. Increased substrate stiffness promotes migration and is controlled by Myosin. Using Drosophila border cell migration as a model of collective cell migration, we identify, for the first time, that the actin bundling protein Fascin limits Myosin activity in vivo. Loss of Fascin results in: increased activated Myosin on the border cells and their substrate, the nurse cells; decreased border cell Myosin dynamics; and increased nurse cell stiffness as measured by atomic force microscopy. Reducing Myosin restores on-time border cell migration in fascin mutant follicles. Further, Fascins actin bundling activity is required to limit Myosin activation. Surprisingly, we find that Fascin regulates Myosin activity in the border cells to control nurse cell stiffness to promote migration. Thus, these data shift the paradigm from a substrate stiffness-centric model of regulating migration, to uncover that collectively migrating cells play a critical role in controlling the mechanical properties of their substrate in order to promote their own migration. This new means of mechanical regulation of migration is likely conserved across contexts and organisms, as Fascin and Myosin are common regulators of cell migration.

cell biology↗

bioRxiv2020

Importin-9 regulates chromosome segregation and packaging in Drosophila germ cells

Germ cells undergo distinct nuclear processes as they differentiate into gametes. While these events must be coordinated to ensure proper maturation, the stage-specific transport of proteins in and out of germ cell nuclei remains incompletely understood. Our efforts to genetically characterize Drosophila genes that exhibit enriched expression in germ cells led to the finding that loss of the highly-conserved Importin {beta}/karyopherin family member Importin-9 (Ipo9) results in female and male sterility. Immunofluorescence and fluorescent in situ hybridization (FISH) revealed that Ipo9KO mutants display chromosome condensation and segregation defects during meiosis. In addition, Ipo9KO mutant males form abnormally structured sperm and fail to properly exchange histones for protamines. Ipo9 physically interacts with proteasome proteins and Ipo9 mutant males exhibit loss of nuclear ubiquitination and disruption of the nuclear localization of several proteasome components. Thus, Ipo9 coordinates the nuclear import of functionally related factors necessary for the completion of gametogenesis. HighlightsDrosophila Importin-9 functions in female and male meiosis Loss of Importin-9 disrupts the histone to protamine transition during spermiogenesis Importin-9 physically interacts with components of the proteasome Importin-9 is required for the efficient nuclear transport of proteasome proteins during spermiogenesis

developmental biology↗

bioRxiv2019

Prostaglandins regulate invasive, collective border cell migration

While prostaglandins (PGs), short-range lipid signals, regulate cell migration, their mechanisms of action are poorly understood in collective migration. To address this, we use Drosophila border cell migration during Stage 9 of oogenesis. The border cells delaminate from the epithelium, and migrate collectively and invasively between the nurse cells. Pxt is the Drosophila cyclooxygenase-like enzyme responsible for all PG synthesis. Loss of Pxt results in both a significant delay in border cell migration during Stage 9 and an increase in cluster length compared to wild-type controls. Contributing to these phenotypes is altered integrin localization. Integrins are enriched on the border cell membranes, and this enrichment is lost in pxt mutants. Active integrins require interaction with the actin cytoskeleton. As we previously found PGs regulate the actin bundler Fascin and Fascin is required for border cell migration, we hypothesized PGs regulate Fascin to control integrins. Supporting this, loss of Fascin results in a pxt-like integrin localization, and dominant genetic interaction studies reveal that co-reduction of Pxt and Fascin results in delayed and elongated border cell clusters. Together these data lead to the model that PG signaling controls Fascin, and thereby integrins, to mediate on-time border cell migration and maintain cluster cohesion.

cell biology↗

bioRxiv2019

Fascin regulates protrusions and delamination to promote invasive, collective cell migration in vivo.

Fascin is an actin bundling protein that is essential for developmental cell migrations and promotes cancer metastasis. In addition to bundling actin, Fascin has several actin-independent roles. Border cell migration during Drosophila oogenesis provides an excellent model to study Fascins various roles during invasive, collective cell migration. Border cell migration requires Fascin. Fascin functions not only within the migrating border cells, but also within the nurse cells, the substrate for this migration. Loss of Fascin results in increased, shorter and mislocalized protrusions during migration. Data supports the model that Fascin promotes the activity of Enabled, an actin elongating factor, to regulate migration. Additionally, loss of Fascin inhibits border cell delamination. These defects are partially due to altered E-cadherin localization in the border cells; this is predicted to be an actin-independent role of Fascin. Overall, Fascin is essential for multiple aspects of this invasive, collective cell migration, and functions in both actin-dependent and -independent manners. These findings have implications beyond Drosophila, as border cell migration has emerged as a model to study mechanisms mediating cancer metastasis.

cell biology↗

bioRxiv2019

Pharmaco-genetic screen to uncover actin regulators targeted by prostaglandins during Drosophila oogenesis

Prostaglandins (PGs) are lipid signaling molecules with numerous physiologic functions, including pain/inflammation, fertility, and cancer. PGs are produced downstream of cyclooxygenase (COX) enzymes, the targets of non-steroidal anti-inflammatory drugs (NSAIDs). In numerous systems, PGs regulate actin cytoskeletal remodeling, however, their mechanisms of action remain largely unknown. To address this deficiency, we undertook a pharmaco-genetic interaction screen during late-stage Drosophila oogenesis. Drosophila oogenesis is as an established model for studying both actin dynamics and PGs. Indeed, during Stage 10B, cage-like arrays of actin bundles surround each nurse cell nucleus, and during Stage 11, the cortical actin contracts, squeezing the cytoplasmic contents into the oocyte. Both of these cytoskeletal properties are required for follicle development and fertility, and are regulated by PGs. Here we describe a pharmaco-genetic interaction screen that takes advantage of the facts that Stage 10B follicles will mature in culture and COX inhibitors, such as aspirin, block this in vitro follicle maturation. In the screen, aspirin was used at a concentration that blocks 50% of the wild-type follicles from maturing in culture. By combining this aspirin treatment with heterozygosity for mutations in actin regulators, we quantitatively identified enhancers and suppressors of COX inhibition. Here we present the screen results and initial follow-up studies on three strong enhancers - Enabled, Capping protein, and non-muscle Myosin II Regulatory Light Chain. Overall, these studies provide new insight into how PGs regulate both actin bundle formation and cellular contraction, properties that are not only essential for development, but are misregulated in diseases.

cell biology↗