scholarly journals Nonmuscle myosin 2 regulates cortical stability during sprouting angiogenesis

2020 ◽  
Vol 31 (18) ◽  
pp. 1974-1987
Author(s):  
Xuefei Ma ◽  
Yutaka Uchida ◽  
Tingyi Wei ◽  
Chengyu Liu ◽  
Ralf H. Adams ◽  
...  
Keyword(s):  
Focal Adhesions ◽  
Nonmuscle Myosin ◽  
Sprouting Angiogenesis ◽  
Vascular Branching

Efficient angiogenic sprouting depends on a dynamic cytoskeleton and focal adhesions. Here we show that nonmuscle myosin 2 functions to promote the retraction of cortical protrusions and the formation of focal adhesions during sprouting, thereby preventing excessive vascular branching.

scholarly journals LIMCH1 regulates nonmuscle myosin-II activity and suppresses cell migration

2017 ◽  
Vol 28 (8) ◽  
pp. 1054-1065 ◽  
Author(s):  
Yu-Hung Lin ◽  
Yen-Yi Zhen ◽  
Kun-Yi Chien ◽  
I-Ching Lee ◽  
Wei-Chi Lin ◽  
...  
Keyword(s):  
Cell Migration ◽  
Hela Cells ◽  
Myosin Ii ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Stress Fibers ◽  
Actin Stress Fiber ◽  
Nonmuscle Myosin ◽  
Head Region ◽  
Nonmuscle Myosin Ii

Nonmuscle myosin II (NM-II) is an important motor protein involved in cell migration. Incorporation of NM-II into actin stress fiber provides a traction force to promote actin retrograde flow and focal adhesion assembly. However, the components involved in regulation of NM-II activity are not well understood. Here we identified a novel actin stress fiber–associated protein, LIM and calponin-homology domains 1 (LIMCH1), which regulates NM-II activity. The recruitment of LIMCH1 into contractile stress fibers revealed its localization complementary to actinin-1. LIMCH1 interacted with NM-IIA, but not NM-IIB, independent of the inhibition of myosin ATPase activity with blebbistatin. Moreover, the N-terminus of LIMCH1 binds to the head region of NM-IIA. Depletion of LIMCH1 attenuated myosin regulatory light chain (MRLC) diphosphorylation in HeLa cells, which was restored by reexpression of small interfering RNA–resistant LIMCH1. In addition, LIMCH1-depleted HeLa cells exhibited a decrease in the number of actin stress fibers and focal adhesions, leading to enhanced cell migration. Collectively, our data suggest that LIMCH1 plays a positive role in regulation of NM-II activity through effects on MRLC during cell migration.

scholarly journals Collagen remodeling by phagocytosis is determined by collagen substrate topology and calcium-dependent interactions of gelsolin with nonmuscle myosin IIA in cell adhesions

2013 ◽  
Vol 24 (6) ◽  
pp. 734-747 ◽  
Author(s):  
P. D. Arora ◽  
Y. Wang ◽  
A. Bresnick ◽  
J. Dawson ◽  
P. A. Janmey ◽  
...  
Keyword(s):  
Focal Adhesions ◽  
Collagen Degradation ◽  
Ion Concentration ◽  
Nonmuscle Myosin ◽  
Collagen Substrate ◽  
Calcium Dependent ◽  
Intracellular Calcium Ion ◽  
Myosin Iia ◽  
Collagen Phagocytosis ◽  
Intracellular Collagen

We examine how collagen substrate topography, free intracellular calcium ion concentration ([Ca2+]i, and the association of gelsolin with nonmuscle myosin IIA (NMMIIA) at collagen adhesions are regulated to enable collagen phagocytosis. Fibroblasts plated on planar, collagen-coated substrates show minimal increase of [Ca2+]i, minimal colocalization of gelsolin and NMMIIA in focal adhesions, and minimal intracellular collagen degradation. In fibroblasts plated on collagen-coated latex beads there are large increases of [Ca2+]i, time- and Ca2+-dependent enrichment of NMMIIA and gelsolin at collagen adhesions, and abundant intracellular collagen degradation. NMMIIA knockdown retards gelsolin recruitment to adhesions and blocks collagen phagocytosis. Gelsolin exhibits tight, Ca2+-dependent binding to full-length NMMIIA. Gelsolin domains G4–G6 selectively require Ca2+ to interact with NMMIIA, which is restricted to residues 1339–1899 of NMMIIA. We conclude that cell adhesion to collagen presented on beads activates Ca2+ entry and promotes the formation of phagosomes enriched with NMMIIA and gelsolin. The Ca2+ -dependent interaction of gelsolin and NMMIIA in turn enables actin remodeling and enhances collagen degradation by phagocytosis.

scholarly journals Endothelial cell invasiveness is controlled by myosin IIA-dependent inhibition of Arp2/3 activity

2020 ◽  
Author(s):  
Ana M. Figueiredo ◽  
Pedro Barbacena ◽  
Ana Russo ◽  
Silvia Vaccaro ◽  
Daniela Ramalho ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Focal Adhesions ◽  
Sprouting Angiogenesis ◽  
Dependent Inhibition ◽  
Myosin Iia ◽  
Maturation State ◽  
Cell Invasiveness ◽  
Cellular Protrusion ◽  
Tip Cells ◽  
New Strategies

AbstractSprouting angiogenesis is fundamental for development and contributes to multiple diseases, including cancer, diabetic retinopathy and cardiovascular diseases. Sprouting angiogenesis depends on the invasive properties of endothelial tip cells. However, there is very limited knowledge on the mechanisms that endothelial tip cells use to invade into tissues. Here, we prove that endothelial tip cells use long lamellipodia projections (LLPs) as the main cellular protrusion for invasion into non-vascular extracellular matrix. We show that LLPs and filopodia protrusions are balanced by myosin-IIA (MIIA) and actin-related protein 2/3 (Arp2/3) activity. Endothelial cell-autonomous ablation of MIIA promotes excessive LLPs formation in detriment of filopodia. Conversely, endothelial cell-autonomous ablation of Arp2/3 prevents LLPs development and leads to excessive filopodia formation. We further show that MIIA inhibits Rac1-dependent activation of Arp2/3, by regulating the maturation state of focal adhesions. Our discoveries establish the first comprehensive model of how endothelial tip cells regulate its protrusive activity and will pave the way towards new strategies to block invasive tip cells during sprouting angiogenesis.

scholarly journals Replacing nonmuscle myosin 2A with myosin 2C1 permits gastrulation but not placenta vascular development in mice

2018 ◽  
Vol 29 (19) ◽  
pp. 2326-2335
Author(s):  
Yingfan Zhang ◽  
Chengyu Liu ◽  
Robert S. Adelstein ◽  
Xuefei Ma
Keyword(s):  
Homologous Recombination ◽  
Focal Adhesions ◽  
Nonmuscle Myosin ◽  
Developmentally Delayed ◽  
Mouse Development ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Heavy Chains ◽  
Vascular Formation

Three paralogues of nonmuscle myosin 2 (NM 2A, 2B, and 2C) are expressed in mammals, and the heavy chains are the products of three different genes (Myh9, Myh10, and Myh14, respectively). NM 2A and 2B are essential for mouse development, while 2C is not. Studies on NM 2C are limited and the in vivo function of this paralogue is not clear. Using homologous recombination, cDNA encoding nonmuscle myosin heavy chain 2C1 fused with GFP was introduced into the first coding exon of Myh9, replacing NM 2A expression with NM 2C1 expression in mice. In contrast to A–/A– embryos, which die by embryonic day (E) 6.5, AC1*gfp/AC1*gfp embryos survive through E8.5, demonstrating that NM 2C1 can support mouse development beyond gastrulation. At E9.5 and E10.5, however, AC1*gfp/AC1*gfp embryos are developmentally delayed, with abnormalities in placental vascular formation. The defect in vascular formation is confirmed in allantois explants from AC1*gfp/AC1*gfp embryos. Thus, NM 2C1 cannot support normal placental vascular formation. In addition, AC1*gfp/AC1*gfp mouse embryonic fibroblasts (MEFs) migrate rapidly but with impaired persistence and develop smaller, less mature focal adhesions than A+/A+ MEFs. This is attributed to enhanced NM 2C1 actomyosin stability and different NM 2C1 subcellular localization than in NM 2A.

scholarly journals Competition for endothelial cell polarity drives vascular morphogenesis

2021 ◽  
Author(s):  
Pedro Barbacena ◽  
Maria Dominguez-Cejudo ◽  
Catarina G. Fonseca ◽  
Manuel Gomez-Gonzalez ◽  
Laura M. Faure ◽  
...  
Keyword(s):  
Shear Stress ◽  
Cell Polarity ◽  
Individual Cell ◽  
Focal Adhesions ◽  
Tissue Level ◽  
Mathematical Methods ◽  
Sprouting Angiogenesis ◽  
Vascular Morphogenesis ◽  
And Migration ◽  
Emergent Behaviors

Blood vessel formation generates unique vascular patterns in each individual. The principles governing the apparent stochasticity of this process remain to be elucidated. Using mathematical methods, we find that the transition between two fundamental vascular morphogenetic programs, sprouting angiogenesis and vascular remodeling, is established by a shift on collective front-rear polarity of endothelial cells. We demonstrate that the competition between biochemical (VEGFA) and mechanical (blood flow-induced shear stress) cues controls this collective polarity shift. Shear stress increases tension at focal adhesions overriding VEGFA-driven collective polarization, which relies on tension at adherens junctions. We propose that vascular morphogenetic cues compete to regulate individual cell polarity and migration through tension shifts that translates into tissue-level emergent behaviors, ultimately leading to uniquely organized vascular patterns.

scholarly journals Conditional Expression of a Truncated Fragment of Nonmuscle Myosin II-A Alters Cell Shape but Not Cytokinesis in HeLa Cells

2000 ◽  
Vol 11 (10) ◽  
pp. 3617-3627 ◽  
Author(s):  
Qize Wei ◽  
Robert S. Adelstein
Keyword(s):  
Hela Cells ◽  
Fusion Proteins ◽  
Myosin Ii ◽  
Focal Adhesions ◽  
Full Length ◽  
Stress Fibers ◽  
Amino Terminus ◽  
Nonmuscle Myosin ◽  
Nonmuscle Myosin Ii ◽  
Conditional Expression

A truncated fragment of the nonmuscle myosin II-A heavy chain (NMHC II-A) lacking amino acids 1–591, ΔN592, was used to examine the cellular functions of this protein. Green fluorescent protein (GFP) was fused to the amino terminus of full-length human NMHC II-A, NMHC II-B, and ΔN592 and the fusion proteins were stably expressed in HeLa cells by using a conditional expression system requiring absence of doxycycline. The HeLa cell line studied normally expressed only NMHC II-A and not NMHC II-B protein. Confocal microscopy indicated that the GFP fusion proteins of full-length NMHC II-A, II-B, and ΔN592 were localized to stress fibers. However, in vitro assays showed that baculovirus-expressed ΔN592 did not bind to actin, suggesting that ΔN592 was localized to actin stress fibers through incorporation into endogenous myosin filaments. There was no evidence for the formation of heterodimers between the full-length endogenous nonmuscle myosin and truncated nonmuscle MHCs. Expression of ΔN592, but not full-length NMHC II-A or NMHC II-B, induced cell rounding with rearrangement of actin filaments and disappearance of focal adhesions. These cells returned to their normal morphology when expression of ΔN592 was repressed by addition of doxycycline. We also show that GFP-tagged full-length NMHC II-A or II-B, but not ΔN592, were localized to the cytokinetic ring during mitosis, indicating that, in vertebrates, the amino-terminus part of mammalian nonmuscle myosin II may be necessary for localization to the cytokinetic ring.

scholarly journals eNOS controls angiogenic sprouting and retinal neovascularization through the regulation of endothelial cell polarity

2021 ◽  
Author(s):  
Tracy L. Smith ◽  
Malika Oubaha ◽  
Gael Cagnone ◽  
Cécile Boscher ◽  
Jin Sung Kim ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Polarity ◽  
Cell Polarization ◽  
No Synthase ◽  
Sprouting Angiogenesis ◽  
Oxygen Induced Retinopathy ◽  
Tip Cells ◽  
Deficient Mice ◽  
Vascular Branching

AbstractThe roles of nitric oxide (NO) and endothelial NO synthase (eNOS) in the regulation of angiogenesis are well documented. However, the involvement of eNOS in the sprouting of endothelial tip-cells at the vascular front during sprouting angiogenesis remains poorly defined. In this study, we show that downregulation of eNOS markedly inhibits VEGF-stimulated migration of endothelial cells but increases their polarization, as evidenced by the reorientation of the Golgi in migrating monolayers and by the fewer filopodia on tip cells at ends of sprouts in endothelial cell spheroids. The effect of eNOS inhibition on EC polarization was prevented in Par3-depleted cells. Importantly, downregulation of eNOS increased the expression of polarity genes, such as PARD3B, PARD6A, PARD6B, PKCΖ, TJP3, and CRB1 in endothelial cells. In retinas of eNOS knockout mice, vascular development is retarded with decreased vessel density and vascular branching. Furthermore, tip cells at the extremities of the vascular front have a marked reduction in the number of filopodia per cell and are more oriented. In a model of oxygen-induced retinopathy (OIR), eNOS deficient mice are protected during the initial vaso-obliterative phase, have reduced pathological neovascularization, and retinal endothelial tip cells have fewer filopodia. Single-cell RNA sequencing of endothelial cells from OIR retinas revealed enrichment of genes related to cell polarity in the endothelial tip-cell subtype of eNOS deficient mice. These results indicate that inhibition of eNOS alters the polarity program of endothelial cells, which increases cell polarization, regulates sprouting angiogenesis and normalizes pathological neovascularization during retinopathy.

scholarly journals Endothelial cell invasion is controlled by dactylopodia

2021 ◽  
Vol 118 (18) ◽  
pp. e2023829118
Author(s):  
Ana Martins Figueiredo ◽  
Pedro Barbacena ◽  
Ana Russo ◽  
Silvia Vaccaro ◽  
Daniela Ramalho ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cell ◽  
Cell Invasion ◽  
Focal Adhesions ◽  
Comprehensive Model ◽  
Related Protein ◽  
Sprouting Angiogenesis ◽  
Maturation State ◽  
Cellular Protrusion ◽  
Tip Cells

Sprouting angiogenesis is fundamental for development and contributes to cancer, diabetic retinopathy, and cardiovascular diseases. Sprouting angiogenesis depends on the invasive properties of endothelial tip cells. However, there is very limited knowledge on how tip cells invade into tissues. Here, we show that endothelial tip cells use dactylopodia as the main cellular protrusion for invasion into nonvascular extracellular matrix. We show that dactylopodia and filopodia protrusions are balanced by myosin IIA (NMIIA) and actin-related protein 2/3 (Arp2/3) activity. Endothelial cell-autonomous ablation of NMIIA promotes excessive dactylopodia formation in detriment of filopodia. Conversely, endothelial cell-autonomous ablation of Arp2/3 prevents dactylopodia development and leads to excessive filopodia formation. We further show that NMIIA inhibits Rac1-dependent activation of Arp2/3 by regulating the maturation state of focal adhesions. Our discoveries establish a comprehensive model of how endothelial tip cells regulate its protrusive activity and will pave the way toward strategies to block invasive tip cells during sprouting angiogenesis.

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