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1.
Cells Dev ; 177: 203883, 2024 03.
Article in English | MEDLINE | ID: mdl-37935283

ABSTRACT

The forces driving tissue morphogenesis are thought to originate from cellular activities. While it is appreciated that extracellular matrix (ECM) may also be involved, ECM function is assumed to be simply instructive in modulating the cellular behaviors that drive changes to tissue shape. However, there is increasing evidence that the ECM may not be the passive player portrayed in developmental biology textbooks. In this review we highlight examples of embryonic ECM dynamics that suggest cell-independent activity, along with developmental processes during which localized ECM alterations and ECM-autonomous forces are directing changes to tissue shape. Additionally, we discuss experimental approaches to unveil active ECM roles during tissue morphogenesis. We propose that it may be time to rethink our general definition of morphogenesis as a cellular-driven phenomenon and incorporate an underappreciated, and surprisingly dynamic ECM.


Subject(s)
Extracellular Matrix , Morphogenesis
2.
Matrix Biol ; 123: 1-16, 2023 Nov.
Article in English | MEDLINE | ID: mdl-37660739

ABSTRACT

Fibrosis is associated with dramatic changes in extracellular matrix (ECM) architecture of unknown etiology. Here we exploit keloid scars as a paradigm to understand fibrotic ECM organization. We reveal that keloid patient fibroblasts uniquely produce a globally aligned ECM network in 2-D culture as observed in scar tissue. ECM anisotropy develops after rapid initiation of a fibroblast supracellular actin network, suggesting that cell alignment initiates ECM patterning. Keloid fibroblasts produce elevated levels of IL-6, and autocrine IL-6 production is both necessary and sufficient to induce cell and ECM alignment, as evidenced by ligand stimulation of normal dermal fibroblasts and treatment of keloid fibroblasts with the function blocking IL-6 receptor monoclonal antibody, tocilizumab. Downstream of IL-6, supracellular organization of keloid fibroblasts is controlled by activation of cell-cell adhesion. Adhesion formation inhibits contact-induced cellular overlap leading to nematic organization of cells and an alignment of focal adhesions. Keloid fibroblasts placed on isotropic ECM align the pre-existing matrix, suggesting that focal adhesion alignment leads to active anisotropic remodeling. These results show that IL-6-induced fibroblast cooperativity can control the development of a nematic ECM, highlighting both IL-6 signaling and cell-cell adhesions as potential therapeutic targets to inhibit this common feature of fibrosis.


Subject(s)
Keloid , Humans , Keloid/drug therapy , Interleukin-6/genetics , Interleukin-6/metabolism , Anisotropy , Cells, Cultured , Extracellular Matrix/metabolism , Fibroblasts/metabolism
3.
Dev Cell ; 58(10): 825-835.e6, 2023 05 22.
Article in English | MEDLINE | ID: mdl-37086718

ABSTRACT

Forces controlling tissue morphogenesis are attributed to cellular-driven activities, and any role for extracellular matrix (ECM) is assumed to be passive. However, all polymer networks, including ECM, can develop autonomous stresses during their assembly. Here, we examine the morphogenetic function of an ECM before reaching homeostatic equilibrium by analyzing de novo ECM assembly during Drosophila ventral nerve cord (VNC) condensation. Asymmetric VNC shortening and a rapid decrease in surface area correlate with the exponential assembly of collagen IV (Col4) surrounding the tissue. Concomitantly, a transient developmentally induced Col4 gradient leads to coherent long-range flow of ECM, which equilibrates the Col4 network. Finite element analysis and perturbation of Col4 network formation through the generation of dominant Col4 mutations that affect assembly reveal that VNC morphodynamics is partially driven by a sudden increase in ECM-driven surface tension. These data suggest that ECM assembly stress and associated network instabilities can actively participate in tissue morphogenesis.


Subject(s)
Drosophila , Extracellular Matrix , Animals , Drosophila/genetics , Extracellular Matrix/physiology , Morphogenesis/physiology , Central Nervous System
4.
Dev Cell ; 56(15): 2137-2139, 2021 08 09.
Article in English | MEDLINE | ID: mdl-34375578

ABSTRACT

Epithelia have an innate yet mysterious capacity to rapidly sense and respond to tissue damage. In this issue of Developmental Cell, O'Connor et al. exploit the genetics of Drosophila to reveal that protease release as a result of tissue injury activates insect cytokines to initiate immediate epithelial repair responses.


Subject(s)
Drosophila , Peptide Hydrolases , Animals , Epithelium
5.
STAR Protoc ; 2(1): 100377, 2021 03 19.
Article in English | MEDLINE | ID: mdl-33786460

ABSTRACT

Protein turnover rate is difficult to obtain experimentally. This protocol shows how to mathematically model turnover rates in an intervention-free manner given the ability to quantify mRNA and protein expression from initiation to homeostasis. This approach can be used to calculate production and degradation rates and to infer protein half-life. This model was successfully employed to quantify turnover during Drosophila embryogenesis, and we hypothesize that it will be applicable to diverse in vivo or in vitro systems. For complete details on the use and execution of this protocol, please refer to Matsubayashi et al. (2020).


Subject(s)
Computational Biology/methods , Proteolysis , RNA, Messenger/metabolism , Animals , Drosophila/metabolism , Gene Expression/genetics , Homeostasis , Kinetics , Models, Theoretical , Proteins/metabolism
6.
Dev Cell ; 54(1): 33-42.e9, 2020 07 06.
Article in English | MEDLINE | ID: mdl-32585131

ABSTRACT

The extracellular matrix (ECM) is a polymer network hypothesized to form a stable cellular scaffold. While the ECM can undergo acute remodeling during embryogenesis, it is experimentally difficult to determine whether basal turnover is also important. Most studies of homeostatic turnover assume an initial steady-state balance of production and degradation and measure half-life by quantifying the rate of decay after experimental intervention (e.g., pulse labeling). Here, we present an intervention-free approach to mathematically model basal ECM turnover during embryogenesis by exploiting our ability to live image de novo ECM development in Drosophila to quantify production from initiation to homeostasis. This reveals rapid turnover (half-life ∼7-10 h), which we confirmed by in vivo pulse-chase experiments. Moreover, ECM turnover is partially dependent on proteolysis and network interactions, and slowing turnover affects tissue morphogenesis. These data demonstrate that embryonic ECM undergoes constant replacement, which is likely necessary to maintain network plasticity to accommodate growth and morphogenesis.


Subject(s)
Extracellular Matrix/metabolism , Homeostasis , Morphogenesis , Animals , Basement Membrane/metabolism , Drosophila Proteins/metabolism , Drosophila melanogaster , Epithelial Cells/cytology , Epithelial Cells/metabolism , Extracellular Matrix Proteins/metabolism , Models, Theoretical
7.
Development ; 147(5)2020 03 02.
Article in English | MEDLINE | ID: mdl-32122911

ABSTRACT

Mutations in the Ultrabithorax (Ubx) gene cause homeotic transformation of the normally two-winged Drosophila into a four-winged mutant fly. Ubx encodes a HOX family transcription factor that specifies segment identity, including transformation of the second set of wings into rudimentary halteres. Ubx is known to control the expression of many genes that regulate tissue growth and patterning, but how it regulates tissue morphogenesis to reshape the wing into a haltere is still unclear. Here, we show that Ubx acts by repressing the expression of two genes in the haltere, Stubble and Notopleural, both of which encode transmembrane proteases that remodel the apical extracellular matrix to promote wing morphogenesis. In addition, Ubx induces expression of the Tissue inhibitor of metalloproteases in the haltere, which prevents the basal extracellular matrix remodelling necessary for wing morphogenesis. Our results provide a long-awaited explanation for how Ubx controls morphogenetic transformation.


Subject(s)
Drosophila Proteins/genetics , Drosophila melanogaster/embryology , Homeodomain Proteins/genetics , Morphogenesis/genetics , Transcription Factors/genetics , Wings, Animal/embryology , Animals , CRISPR-Cas Systems , Drosophila melanogaster/genetics , Matrix Metalloproteinase Inhibitors/metabolism , Membrane Proteins/genetics , Serine Endopeptidases/genetics
8.
Cells ; 9(1)2020 01 08.
Article in English | MEDLINE | ID: mdl-31936297

ABSTRACT

In order to ascertain their external environment, cells and tissues have the capability to sense and process a variety of stresses, including stretching and compression forces. These mechanical forces, as experienced by cells and tissues, are then converted into biochemical signals within the cell, leading to a number of cellular mechanisms being activated, including proliferation, differentiation and migration. If the conversion of mechanical cues into biochemical signals is perturbed in any way, then this can be potentially implicated in chronic disease development and processes such as neurological disorders, cancer and obesity. This review will focus on how the interplay between mechanotransduction, cellular structure, metabolism and signalling cascades led by the Hippo-YAP/TAZ axis can lead to a number of chronic diseases and suggest how we can target various pathways in order to design therapeutic targets for these debilitating diseases and conditions.


Subject(s)
Cell Cycle Proteins/metabolism , Chronic Disease/epidemiology , Mechanotransduction, Cellular , Protein Serine-Threonine Kinases/metabolism , Transcription Factors/metabolism , Acyltransferases , Hippo Signaling Pathway , Humans , Signal Transduction
9.
J Cell Sci ; 131(22)2018 11 22.
Article in English | MEDLINE | ID: mdl-30404826

ABSTRACT

Human cells can sense mechanical stress acting upon integrin adhesions and respond by sending the YAP (also known as YAP1) and TAZ (also known as WWTR1) transcriptional co-activators to the nucleus to drive TEAD-dependent transcription of target genes. How integrin signaling activates YAP remains unclear. Here, we show that integrin-mediated mechanotransduction requires the Enigma and Enigma-like proteins (PDLIM7 and PDLIM5, respectively; denoted for the family of PDZ and LIM domain-containing proteins). YAP binds to PDLIM5 and PDLIM7 (hereafter PDLIM5/7) via its C-terminal PDZ-binding motif (PBM), which is essential for full nuclear localization and activity of YAP. Accordingly, silencing of PDLIM5/7 expression reduces YAP nuclear localization, tyrosine phosphorylation and transcriptional activity. The PDLIM5/7 proteins are recruited from the cytoplasm to integrin adhesions and F-actin stress fibers in response to force by binding directly to the key stress fiber component α-actinin. Thus, forces acting on integrins recruit Enigma family proteins to trigger YAP activation during mechanotransduction.This article has an associated First Person interview with the first author of the paper.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Cytoskeletal Proteins/metabolism , LIM Domain Proteins/metabolism , Transcription Factors/metabolism , Animals , Caco-2 Cells , Fibroblasts/metabolism , HEK293 Cells , Humans , Integrins/metabolism , Mechanotransduction, Cellular , Mice , Signal Transduction , Trans-Activators , Transcriptional Coactivator with PDZ-Binding Motif Proteins , YAP-Signaling Proteins
10.
Dev Cell ; 46(1): 23-39.e5, 2018 07 02.
Article in English | MEDLINE | ID: mdl-29974861

ABSTRACT

Epithelial tissues can elongate in two dimensions by polarized cell intercalation, oriented cell division, or cell shape change, owing to local or global actomyosin contractile forces acting in the plane of the tissue. In addition, epithelia can undergo morphogenetic change in three dimensions. We show that elongation of the wings and legs of Drosophila involves a columnar-to-cuboidal cell shape change that reduces cell height and expands cell width. Remodeling of the apical extracellular matrix by the Stubble protease and basal matrix by MMP1/2 proteases induces wing and leg elongation. Matrix remodeling does not occur in the haltere, a limb that fails to elongate. Limb elongation is made anisotropic by planar polarized Myosin-II, which drives convergent extension along the proximal-distal axis. Subsequently, Myosin-II relocalizes to lateral membranes to accelerate columnar-to-cuboidal transition and isotropic tissue expansion. Thus, matrix remodeling induces dynamic changes in actomyosin contractility to drive epithelial morphogenesis in three dimensions.


Subject(s)
Body Patterning/physiology , Drosophila melanogaster/embryology , Epithelial Cells/cytology , Lower Extremity/embryology , Morphogenesis/physiology , Wings, Animal/embryology , Animals , Cell Polarity/physiology , Cell Shape/physiology , Drosophila Proteins/metabolism , Drosophila melanogaster/cytology , Embryo, Nonmammalian/embryology , Epithelium/metabolism , Matrix Metalloproteinase 1/metabolism , Matrix Metalloproteinase 2/metabolism , Membrane Proteins/metabolism , Myosin Type II/metabolism , Serine Endopeptidases/metabolism
11.
Development ; 145(5)2018 03 08.
Article in English | MEDLINE | ID: mdl-29440303

ABSTRACT

Animal cells are thought to sense mechanical forces via the transcriptional co-activators YAP (or YAP1) and TAZ (or WWTR1), the sole Drosophila homolog of which is named Yorkie (Yki). In mammalian cells in culture, artificial mechanical forces induce nuclear translocation of YAP and TAZ. Here, we show that physiological mechanical strain can also drive nuclear localisation of Yki and activation of Yki target genes in the Drosophila follicular epithelium. Mechanical strain activates Yki by stretching the apical domain, reducing the concentration of apical Crumbs, Expanded, Kibra and Merlin, and reducing apical Hippo kinase dimerisation. Overexpressing Hippo kinase to induce ectopic activation in the cytoplasm is sufficient to prevent Yki nuclear localisation even in flattened follicle cells. Conversely, blocking Hippo signalling in warts clones causes Yki nuclear localisation even in columnar follicle cells. We find no evidence for involvement of other pathways, such as Src42A kinase, in regulation of Yki. Finally, our results in follicle cells appear generally applicable to other tissues, as nuclear translocation of Yki is also readily detectable in other flattened epithelial cells such as the peripodial epithelium of the wing imaginal disc, where it promotes cell flattening.


Subject(s)
Drosophila Proteins/metabolism , Drosophila melanogaster , Intracellular Signaling Peptides and Proteins/metabolism , Protein Serine-Threonine Kinases/metabolism , Stress, Mechanical , Wings, Animal/embryology , Animals , Animals, Genetically Modified , Body Patterning/genetics , Cell Nucleus/metabolism , Drosophila Proteins/genetics , Drosophila melanogaster/embryology , Drosophila melanogaster/genetics , Drosophila melanogaster/metabolism , Embryo, Nonmammalian , Gene Expression Regulation, Developmental , Imaginal Discs/embryology , Imaginal Discs/metabolism , Intracellular Signaling Peptides and Proteins/genetics , Mechanotransduction, Cellular/physiology , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Protein Serine-Threonine Kinases/genetics , Protein Transport/genetics , Signal Transduction/genetics , Trans-Activators/genetics , Trans-Activators/metabolism , Transcription Factors/metabolism , Wings, Animal/metabolism , YAP-Signaling Proteins
13.
J Cell Sci ; 129(13): 2651-9, 2016 07 01.
Article in English | MEDLINE | ID: mdl-27231092

ABSTRACT

In epithelial tissues, polarisation of microtubules and actin microvilli occurs along the apical-basal axis of each cell, yet how these cytoskeletal polarisation events are coordinated remains unclear. Here, we examine the hierarchy of events during cytoskeletal polarisation in Drosophila melanogaster epithelia. Core apical-basal polarity determinants polarise the spectrin cytoskeleton to recruit the microtubule-binding proteins Patronin (CAMSAP1, CAMSAP2 and CAMSAP3 in humans) and Shortstop [Shot; MACF1 and BPAG1 (also known as DST) in humans] to the apical membrane domain. Patronin and Shot then act to polarise microtubules along the apical-basal axis to enable apical transport of Rab11 endosomes by the Nuf-Dynein microtubule motor complex. Finally, Rab11 endosomes are transferred to the MyoV (also known as Didum in Drosophila) actin motor to deliver the key microvillar determinant Cadherin 99C to the apical membrane to organise the biogenesis of actin microvilli.


Subject(s)
Drosophila Proteins/genetics , Microfilament Proteins/genetics , Microtubule-Associated Proteins/genetics , Microvilli/metabolism , Myosin Type V/genetics , rab GTP-Binding Proteins/genetics , Actin Cytoskeleton/genetics , Actin Cytoskeleton/metabolism , Animals , Cadherins/metabolism , Cell Membrane/genetics , Cell Membrane/metabolism , Cell Polarity/genetics , Drosophila Proteins/metabolism , Drosophila melanogaster/genetics , Epithelium/growth & development , Epithelium/metabolism , Humans , Microtubules/genetics , Microvilli/genetics , Myosin Type V/metabolism , Protein Transport/genetics , rab GTP-Binding Proteins/metabolism
14.
PLoS Genet ; 12(1): e1005763, 2016 Jan.
Article in English | MEDLINE | ID: mdl-26808525

ABSTRACT

The extracellular matrix (ECM) is a pivotal component adult tissues and of many tissue-specific stem cell niches. It provides structural support and regulates niche signaling during tissue maintenance and regeneration. In many tissues, ECM remodeling depends on the regulation of MMP (matrix metalloproteinase) activity by inhibitory TIMP (tissue inhibitors of metalloproteinases) proteins. Here, we report that the only Drosophila timp gene is required for maintaining the normal organization and function of the germline stem cell niche in adult females. timp mutant ovaries show reduced levels of both Drosophila Collagen IV α chains. In addition, tissue stiffness and the cellular organization of the ovarian niche are affected in timp mutants. Finally, loss of timp impairs the ability of the germline stem cell niche to generate new cysts. Our results demonstrating a crucial role for timp in tissue organization and gamete production thus provide a link between the regulation of ECM metabolism and tissue homeostasis.


Subject(s)
Extracellular Matrix/metabolism , Ovary/metabolism , Stem Cell Niche/genetics , Tissue Inhibitor of Metalloproteinases/genetics , Animals , Collagen Type IV/genetics , Drosophila , Extracellular Matrix/genetics , Female , Germ Cells , Matrix Metalloproteinases/genetics , Ovary/growth & development
15.
J Biol Chem ; 287(22): 18717-29, 2012 May 25.
Article in English | MEDLINE | ID: mdl-22493290

ABSTRACT

Manganese is an essential trace element, whose intracellular levels need to be carefully regulated. Mn(2+) acts as a cofactor for many enzymes and excess of Mn(2+) is toxic. Alterations in Mn(2+) homeostasis affect metabolic functions and mutations in the human Mn(2+)/Ca(2+) transporter ATP2C1 have been linked to Hailey-Hailey disease. By deletion of the yeast orthologue PMR1 we have studied the impact of Mn(2+) on cell cycle progression and show that an excess of cytosolic Mn(2+) alters S-phase transit, induces transcriptional up-regulation of cell cycle regulators, bypasses the need for S-phase cell cycle checkpoints and predisposes to genomic instability. On the other hand, we find that depletion of the Golgi Mn(2+) pool requires a functional morphology checkpoint to avoid the formation of polyploid cells.


Subject(s)
Manganese/metabolism , Mitosis , Blotting, Western , Cell Cycle , Flow Cytometry , Genomic Instability , Homeostasis
16.
Nucleic Acids Res ; 39(14): 6002-15, 2011 Aug.
Article in English | MEDLINE | ID: mdl-21511814

ABSTRACT

Genomic instability is related to a wide-range of human diseases. Here, we show that mitochondrial iron-sulfur cluster biosynthesis is important for the maintenance of nuclear genome stability in Saccharomyces cerevisiae. Cells lacking the mitochondrial chaperone Zim17 (Tim15/Hep1), a component of the iron-sulfur biosynthesis machinery, have limited respiration activity, mimic the metabolic response to iron starvation and suffer a dramatic increase in nuclear genome recombination. Increased oxidative damage or deficient DNA repair do not account for the observed genomic hyperrecombination. Impaired cell-cycle progression and genetic interactions of ZIM17 with components of the RFC-like complex involved in mitotic checkpoints indicate that replicative stress causes hyperrecombination in zim17Δ mutants. Furthermore, nuclear accumulation of pre-ribosomal particles in zim17Δ mutants reinforces the importance of iron-sulfur clusters in normal ribosome biosynthesis. We propose that compromised ribosome biosynthesis and cell-cycle progression are interconnected, together contributing to replicative stress and nuclear genome instability in zim17Δ mutants.


Subject(s)
Cell Nucleus/genetics , Genomic Instability , Iron-Sulfur Proteins/biosynthesis , Mitochondrial Proteins/physiology , Saccharomyces cerevisiae Proteins/physiology , DNA Damage , DNA Replication , Gene Deletion , Gene Expression Regulation, Fungal , Iron/metabolism , Mitochondrial Proteins/genetics , Mutation , Recombinases/metabolism , Recombination, Genetic , Replication Protein C/metabolism , Ribosomes/metabolism , S Phase , Saccharomyces cerevisiae/enzymology , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/genetics , Transcription, Genetic
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