RhoA-ROCK competes with YAP to regulate amoeboid breast cancer cell migration in response to lymphatic-like flow
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA789864 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR17262774_1.fastq.gz | Fastqsanger.gz | |||
| SRR17262774_2.fastq.gz | Fastqsanger.gz | |||
| SRR17262775_1.fastq.gz | Fastqsanger.gz | |||
| SRR17262775_2.fastq.gz | Fastqsanger.gz | |||
| SRR17262776_1.fastq.gz | Fastqsanger.gz |
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Project description:Lymphatic drainage generates force that induces prostate cancer cell motility via activation of Yes-associated protein (YAP), but whether this response to fluid force is conserved across cancer types is unclear. Here, we show that shear stress corresponding to fluid flow in the initial lymphatics modifies taxis in breast cancer. Whereas some cell lines employ rapid amoeboid migration behavior in response to fluid flow, a separate subset decrease movement. Positive responders displayed transcriptional profiles typical of an amoeboid cell state. Regulation of the HIPPO tumor suppressor pathway and YAP activity also differed between breast subsets and prostate cancer. Although subcellular localization of YAP to the nucleus positively correlated with overall velocity of locomotion, YAP gain- and loss-of-function demonstrates that YAP inhibits breast cancer motility but is outcompeted by other pro-taxis mediators in the context of flow. Specifically, we show that RhoA dictates response to flow. GTPase activity of RhoA, but not Rac1 or Cdc42 Rho family GTPases, is elevated in cells that positively respond to flow and is unchanged in cells that decelerate under flow. Disruption of RhoA or the RhoA effector, Rho-associated kinase (ROCK), blocked shear stress-induced motility. Collectively, these findings demonstrate stratification of breast cancer subsets by flow-sensing mechanotransduction pathways and point to a role for biophysical force in regulation of an amoeboid cell state.
2022-03-27 | GSE191142 | GEO
Project description:Lymphatic drainage generates force that induces prostate cancer cell motility via activation of Yes-associated protein (YAP), but whether this response to fluid force is conserved across cancer types is unclear. Here, we show that shear stress corresponding to fluid flow in the initial lymphatics modifies taxis in breast cancer, whereas some cell lines use rapid amoeboid migration behavior in response to fluid flow, a separate subset decrease movement. Positive responders displayed transcriptional profiles characteristic of an amoeboid cell state, which is typical of cells advancing at the edges of neoplastic tumors. Regulation of the HIPPO tumor suppressor pathway and YAP activity also differed between breast subsets and prostate cancer. Although subcellular localization of YAP to the nucleus positively correlated with overall velocity of locomotion, YAP gain- and loss-of-function demonstrates that YAP inhibits breast cancer motility but is outcompeted by other pro-taxis mediators in the context of flow. Specifically, we show that RhoA dictates response to flow. GTPase activity of RhoA, but not Rac1 or Cdc42 Rho family GTPases, is elevated in cells that positively respond to flow and is unchanged in cells that decelerate under flow. Disruption of RhoA or the RhoA effector, Rho-associated kinase (ROCK), blocked shear stress-induced motility. Collectively, these findings identify biomechanical force as a regulator amoeboid cell migration and demonstrate stratification of breast cancer subsets by flow-sensing mechanotransduction pathways.
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Project description:We have discovered a previously uncharacterized tumor suppressor function of Numb in the homeostasis of the normal bladder mucosa. Targeted ablation of Numb in the basal layer of the urothelium drives spontaneous bladder tumorigenesis, driving progression from preneoplastic to preinvasive and overtly invasive tumors, and accelerates tumorigenesis in the presence of oncogenic insults. Using 3D-Matrigel organoid cultures to recapitulate bladder tumorigenesis in vitro, we found that Numb deficiency enhances the proliferative and invasive potential of both mouse and human bladder cancer cells. Integrative transcriptomic and functional analyses revealed that downregulation of the canonical Hippo pathway, resulting in enhanced YAP transcriptional activity, underlies the biological aggressiveness of Numb-deficient bladder cancer. These molecular events are dependent on the activation of RhoA/ROCK signaling subsequent to Numb loss. Thus, a dysfunctional Numb–RhoA/ROCK–Hippo/YAP regulatory network is at play in aggressive Numb-deficient bladder cancers, opening avenues for the development of targeted therapies. Furthermore, retrospective cohort studies revealed that a Numb-deficient status correlates with worse overall survival in post-cystectomy muscle-invasive bladder cancer (MIBC) patients and increased risk of progression to MIBC in non-muscle-invasive bladder cancer (NMIBC) patients. The Numb-deficient status was associated with a 27-gene prognostic signature that has the potential to identify high-risk NMIBC patients who are eligible for targeted RhoA/ROCK/YAP therapies.
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Project description:During development, multicellular rosettes serve as important cellular intermediates in the formation of diverse organ systems. Multicellular rosettes are transient epithelial structures that are defined by the apical constriction of cells towards the rosette center. Due to the important role these structures play during development, understanding the molecular mechanisms by which rosettes are formed and maintained is of high interest. Utilizing the zebrafish posterior lateral line primordium (pLLP) as a model system, we identify the RhoA GEF Mcf2lb as a regulator of rosette integrity. The pLLP is a group of ~150 cells that migrates along the zebrafish trunk and is organized into epithelial rosettes; these are deposited along the trunk and will differentiate into sensory organs called neuromasts (NMs). Using single-cell RNA sequencing and whole-mount in situ hybridization, we showed that mcf2lb is expressed in the pLLP during migration. Given the known role of RhoA in rosette formation, we asked whether Mcf2lb plays a role in regulating apical constriction of cells within rosettes. Live imaging and subsequent 3D analysis of mcf2lb mutant pLLP cells showed disrupted apical constriction and subsequent rosette organization. This in turn resulted in a unique posterior Lateral Line phenotype: an excess number of deposited NMs along the trunk of the zebrafish. Cell polarity markers ZO-1 and Par-3 were apically localized, indicating that pLLP cells are normally polarized. In contrast, signaling components that mediate apical constriction downstream of RhoA, Rock-2a and non-muscle Myosin II were diminished apically. Altogether our results suggest a model whereby Mcf2lb activates RhoA, which in turn activates downstream signaling machinery to induce and maintain apical constriction in cells incorporated into rosettes.
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Project description:To identify novel proteins that control melanoma development and progression, we integrated high-throughput proteomics, mRNA and miRNAs datasets from isogenic primary (WM115) and metastatic (WM266-4) melanoma cell-lines. We focused on differentially expressed proteins that had non-differential mRNA levels and were putatively targeted by a high number of miRNAs. SNX18, a sub-family member of sorting-nexins that participates in endocytosis and autophagy was down-regulated by 6-fold in WM266-4, and predicted to be targeted by 17 miRNAs. Over-expression of SNX18 in several melanoma cell-lines significantly reduced migration rate, while silencing of SNX18 resulted in the opposite effect. Notably, progression tissue microarrays demonstrated a trend for SNX18 downregulation in early metastasis. Luciferase assays confirmed the direct regulation of SNX18 by multiple miRNAs. Indeed, over-expression of these miRNAs repressed endogenous SNX18 expression at the protein but not mRNA level. Further, protein but not mRNA expression levels correlated with better overall survival as determined in a cohort of 48 metastatic melanoma patients and in the TCGA database. Mechanistically, SNX18-mediated reduced migration coincided with RhoA activation and Cdc42 inactivation. Both play a role, as RhoA inhibitor and constitutively activated Cdc42 mutants abrogated this effect. Moreover, phosphorylation-defective SNX18 mutants (S233A,S233D) migrated faster, with S233A inactivating RhoA and activating Cdc42, while S233D activated both. The downstream phosphorylation of the p38>MAPKAPK2>HSP27 signaling pathway was concordant. Both mutants were more abundant in the plasma membrane. Taken together, phosphorylation of S233 is crucial for SNX18 activation, but de-phosphorylation dynamics, which may occur at the membrane, is required for functional effect. miRNA expression profile of primary (WM115) and metastatic (WM266-4) melanoma cell lines.
2021-09-17 | GSE120033 | GEO
Project description:To identify novel proteins that control melanoma development and progression, we integrated high-throughput proteomics, mRNA and miRNAs datasets from isogenic primary (WM115) and metastatic (WM266-4) melanoma cell-lines. We focused on differentially expressed proteins that had non-differential mRNA levels and were putatively targeted by a high number of miRNAs. SNX18, a sub-family member of sorting-nexins that participates in endocytosis and autophagy was down-regulated by 6-fold in WM266-4, and predicted to be targeted by 17 miRNAs. Over-expression of SNX18 in several melanoma cell-lines significantly reduced migration rate, while silencing of SNX18 resulted in the opposite effect. Notably, progression tissue microarrays demonstrated a trend for SNX18 downregulation in early metastasis. Luciferase assays confirmed the direct regulation of SNX18 by multiple miRNAs. Indeed, over-expression of these miRNAs repressed endogenous SNX18 expression at the protein but not mRNA level. Further, protein but not mRNA expression levels correlated with better overall survival as determined in a cohort of 48 metastatic melanoma patients and in the TCGA database. Mechanistically, SNX18-mediated reduced migration coincided with RhoA activation and Cdc42 inactivation. Both play a role, as RhoA inhibitor and constitutively activated Cdc42 mutants abrogated this effect. Moreover, phosphorylation-defective SNX18 mutants (S233A,S233D) migrated faster, with S233A inactivating RhoA and activating Cdc42, while S233D activated both. The downstream phosphorylation of the p38>MAPKAPK2>HSP27 signaling pathway was concordant. Both mutants were more abundant in the plasma membrane. Taken together, phosphorylation of S233 is crucial for SNX18 activation, but de-phosphorylation dynamics, which may occur at the membrane, is required for functional effect. In this study we compared the proteome, transcriptome and miRnome of primary and metastatic melanoma isogenic cell-lines and identified SNX18 as a novel inhibitor of melanoma migration.
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Project description:To identify novel proteins that control melanoma development and progression, we integrated high-throughput proteomics, mRNA and miRNAs datasets from isogenic primary (WM115) and metastatic (WM266-4) melanoma cell-lines. We focused on differentially expressed proteins that had non-differential mRNA levels and were putatively targeted by a high number of miRNAs. SNX18, a sub-family member of sorting-nexins that participates in endocytosis and autophagy was down-regulated by 6-fold in WM266-4, and predicted to be targeted by 17 miRNAs. Over-expression of SNX18 in several melanoma cell-lines significantly reduced migration rate, while silencing of SNX18 resulted in the opposite effect. Notably, progression tissue microarrays demonstrated a trend for SNX18 downregulation in early metastasis. Luciferase assays confirmed the direct regulation of SNX18 by multiple miRNAs. Indeed, over-expression of these miRNAs repressed endogenous SNX18 expression at the protein but not mRNA level. Further, protein but not mRNA expression levels correlated with better overall survival as determined in a cohort of 48 metastatic melanoma patients and in the TCGA database. Mechanistically, SNX18-mediated reduced migration coincided with RhoA activation and Cdc42 inactivation. Both play a role, as RhoA inhibitor and constitutively activated Cdc42 mutants abrogated this effect. Moreover, phosphorylation-defective SNX18 mutants (S233A,S233D) migrated faster, with S233A inactivating RhoA and activating Cdc42, while S233D activated both. The downstream phosphorylation of the p38>MAPKAPK2>HSP27 signaling pathway was concordant. Both mutants were more abundant in the plasma membrane. Taken together, phosphorylation of S233 is crucial for SNX18 activation, but de-phosphorylation dynamics, which may occur at the membrane, is required for functional effect. mRNA expression profile of primary (WM115) and metastatic (WM266-4) melanoma cell lines.
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2008-10-26 | E-GEOD-12917 | biostudies-arrayexpress
Project description:Collective migration plays critical roles in developmental, physiological and pathological processes, and requires a dynamic actomyosin network for cell shape change, cell adhesion and cell-cell communication. The dynamic network of mitochondria in individual cells is regulated by mitochondrial fission and fusion, and is required for cellular processes including cell metabolism, apoptosis and cell division. But whether mitochondrial dynamics interplays with and regulates actomyosin dynamics during collective migration is not clear. Here, we demonstrate that proper regulation of mitochondrial dynamics is critical for collective migration of Drosophila border cells during oogenesis, and misregulation of fission or fusion results in reduction of ATP levels. Specifically, Drp1 is genetically required for border cell migration, and Drp1-mediated mitochondrial fission promotes formation of leading protrusion, likely through its regulation of ATP levels. Reduction of ATP levels by drug treatment also affects protrusion formation as well as actomyosin dynamics. Importantly, we find that RhoA/ROCK signaling, which is essential for actin and myosin dynamics during border cell migration, could exert its effect on mitochondrial fission through regulating Drp1's recruitment to mitochondria. These findings suggest that RhoA/ROCK signaling may couple or coordinate actomyosin dynamics with mitochondrial dynamics to achieve optimal actomyosin function, leading to protrusive and migratory behavior.
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Project description:Using mice with a keratinocyte-restricted deletion of the RhoA gene, we explored its role in the regulation of gene expression in vivo and in vitro RNA was isolated from RhoA-null or wildtype keratinocytes, in two different conditions, freshly isolated (in vivo) or cultured (in vitro), using the RNeasy kit (Qiagen, Germany). Hybridization to GeneChip Mouse Genome 430 2.0 Array (Affymetrix, Santa Clara, CA) was carried out at the Copenhagen University Hospital Microarray Center using standard protocols. 2 biological replicates were analyzed for each in vivo genotype (2 samples from RhoA-null, 2 from wildtype), while 3 replicates for each in vitro genotype.
2015-01-07 | E-GEOD-64714 | biostudies-arrayexpress