scholarly journals F-actin bundles direct the initiation and orientation of lamellipodia through adhesion-based signaling

2015 ◽  
Vol 208 (4) ◽  
pp. 443-455 ◽  
Author(s):  
Heath E. Johnson ◽  
Samantha J. King ◽  
Sreeja B. Asokan ◽  
Jeremy D. Rotty ◽  
James E. Bear ◽  
...  
Keyword(s):  
Focal Adhesion ◽  
Total Internal Reflection ◽  
Cell Shape ◽  
Platelet Derived Growth Factor ◽  
Pi3k Signaling ◽  
Actin Bundles ◽  
Signaling Dynamics ◽  
Ratiometric Imaging ◽  
Phosphoinositide 3 Kinase

Mesenchymal cells such as fibroblasts are weakly polarized and reorient directionality by a lamellipodial branching mechanism that is stabilized by phosphoinositide 3-kinase (PI3K) signaling. However, the mechanisms by which new lamellipodia are initiated and directed are unknown. Using total internal reflection fluorescence microscopy to monitor cytoskeletal and signaling dynamics in migrating cells, we show that peripheral F-actin bundles/filopodia containing fascin-1 serve as templates for formation and orientation of lamellipodia. Accordingly, modulation of fascin-1 expression tunes cell shape, quantified as the number of morphological extensions. Ratiometric imaging reveals that F-actin bundles/filopodia play both structural and signaling roles, as they prime the activation of PI3K signaling mediated by integrins and focal adhesion kinase. Depletion of fascin-1 ablated fibroblast haptotaxis on fibronectin but not platelet-derived growth factor chemotaxis. Based on these findings, we conceptualize haptotactic sensing as an exploration, with F-actin bundles directing and lamellipodia propagating the process and with signaling mediated by adhesions playing the role of integrator.

scholarly journals Migrating fibroblasts reorient directionality by a metastable, PI3K-dependent mechanism

2012 ◽  
Vol 197 (1) ◽  
pp. 105-114 ◽  
Author(s):  
Erik S. Welf ◽  
Shoeb Ahmed ◽  
Heath E. Johnson ◽  
Adam T. Melvin ◽  
Jason M. Haugh
Keyword(s):  
Cell Migration ◽  
Total Internal Reflection ◽  
Mesenchymal Cell ◽  
Lateral Spreading ◽  
Pi3k Signaling ◽  
Spatial Cues ◽  
Cell Protrusion ◽  
Pi3k Inhibition ◽  
Phosphoinositide 3 Kinase ◽  
Dependent Mechanism

Mesenchymal cell migration as exhibited by fibroblasts is distinct from amoeboid cell migration and is characterized by dynamic competition among multiple protrusions, which determines directional persistence and responses to spatial cues. Localization of phosphoinositide 3-kinase (PI3K) signaling is thought to play a broadly important role in cell motility, yet the context-dependent functions of this pathway have not been adequately elucidated. By mapping the spatiotemporal dynamics of cell protrusion/retraction and PI3K signaling monitored by total internal reflection fluorescence microscopy, we show that randomly migrating fibroblasts reorient polarity through PI3K-dependent branching and pivoting of protrusions. PI3K inhibition did not affect the initiation of newly branched protrusions, nor did it prevent protrusion induced by photoactivation of Rac. Rather, PI3K signaling increased after, not before, the onset of local protrusion and was required for the lateral spreading and stabilization of nascent branches. During chemotaxis, the branch experiencing the higher chemoattractant concentration was favored, and, thus, the cell reoriented so as to align with the external gradient.

scholarly journals Reciprocal regulation among TRPV1 channels and phosphoi nos itide 3-kinase in response to nerve growth factor

2018 ◽  
Author(s):  
Anastasiia Stratiievska ◽  
Sara Nelson ◽  
Eric N. Senning ◽  
Jonathan D. Lautz ◽  
Stephen E.P. Smith ◽  
...  
Keyword(s):  
Total Internal Reflection ◽  
Inflammatory Mediator ◽  
Trp Channels ◽  
Pi3k Signaling ◽  
Reciprocal Regulation ◽  
Trpv1 Channels ◽  
Pi3k Activity ◽  
Trpv Channels ◽  
Receptor Neurons ◽  
Phosphoinositide 3 Kinase

AbstractAlthough it has been known for over a decade that the inflammatory mediator NGF sensitizes pain-receptor neurons through increased trafficking of TRPV1 channels to the plasma membrane, the mechanism by which this occurs remains mysterious. NGF activates phosphoinositide 3-kinase (PI3K), the enzyme that generates PIP3, and PI3K activity is required for sensitization. One tantalizing hint came from the finding that the N-terminal region of TRPV1 interacts directly with PI3K. Using 2-color total internal reflection fluorescence microscopy, we show that TRPV1 potentiates NGF-induced PI3K activity. A soluble TRPV1 fragment corresponding to the N-terminal Ankyrin repeats domain (ARD) was sufficient to produce this potentiation, indicating that allosteric regulation was involved. Further, other TRPV channels with conserved ARDs also potentiated NGF-induced PI3K activity whereas TRP channels lacking ARDs did not. Our data demonstrate a novel reciprocal regulation of PI3K signaling by the ARD of TRPV channels.

scholarly journals Quantitative elucidation of a distinct spatial gradient-sensing mechanism in fibroblasts

2005 ◽  
Vol 171 (5) ◽  
pp. 883-892 ◽  
Author(s):  
Ian C. Schneider ◽  
Jason M. Haugh
Keyword(s):  
Signal Amplification ◽  
Total Internal Reflection ◽  
Feedback Loops ◽  
Platelet Derived Growth Factor ◽  
Spatial Gradient ◽  
Gradient Sensing ◽  
Fibroblast Migration ◽  
Phosphoinositide 3 Kinase ◽  
Kinase Pathway ◽  
Amoeboid Cells

Migration of eukaryotic cells toward a chemoattractant often relies on their ability to distinguish receptor-mediated signaling at different subcellular locations, a phenomenon known as spatial sensing. A prominent example that is seen during wound healing is fibroblast migration in platelet-derived growth factor (PDGF) gradients. As in the well-characterized chemotactic cells Dictyostelium discoideum and neutrophils, signaling to the cytoskeleton via the phosphoinositide 3-kinase pathway in fibroblasts is spatially polarized by a PDGF gradient; however, the sensitivity of this process and how it is regulated are unknown. Through a quantitative analysis of mathematical models and live cell total internal reflection fluorescence microscopy experiments, we demonstrate that PDGF detection is governed by mechanisms that are fundamentally different from those in D. discoideum and neutrophils. Robust PDGF sensing requires steeper gradients and a much narrower range of absolute chemoattractant concentration, which is consistent with a simpler system lacking the feedback loops that yield signal amplification and adaptation in amoeboid cells.

scholarly journals Postnatal tendon growth and remodeling require platelet-derived growth factor receptor signaling

2018 ◽  
Vol 314 (4) ◽  
pp. C389-C403 ◽  
Author(s):  
Kristoffer B. Sugg ◽  
James F. Markworth ◽  
Nathaniel P. Disser ◽  
Andrew M. Rizzi ◽  
Jeffrey R. Talarek ◽  
...  
Keyword(s):  
Growth Factor ◽  
Biological Activities ◽  
Growth Factor Receptor ◽  
Normal Growth ◽  
Platelet Derived Growth Factor ◽  
Growth And Remodeling ◽  
Fibroblast Migration ◽  
And Migration ◽  
Phosphoinositide 3 Kinase

Platelet-derived growth factor receptor (PDGFR) signaling plays an important role in the fundamental biological activities of many cells that compose musculoskeletal tissues. However, little is known about the role of PDGFR signaling during tendon growth and remodeling in adult animals. Using the hindlimb synergist ablation model of tendon growth, our objectives were to determine the role of PDGFR signaling in the adaptation of tendons subjected to a mechanical growth stimulus, as well as to investigate the biological mechanisms behind this response. We demonstrate that both PDGFRs, PDGFRα and PDGFRβ, are expressed in tendon fibroblasts and that the inhibition of PDGFR signaling suppresses the normal growth of tendon tissue in response to mechanical growth cues due to defects in fibroblast proliferation and migration. We also identify membrane type-1 matrix metalloproteinase (MT1-MMP) as an essential proteinase for the migration of tendon fibroblasts through their extracellular matrix. Furthermore, we report that MT1-MMP translation is regulated by phosphoinositide 3-kinase/Akt signaling, while ERK1/2 controls posttranslational trafficking of MT1-MMP to the plasma membrane of tendon fibroblasts. Taken together, these findings demonstrate that PDGFR signaling is necessary for postnatal tendon growth and remodeling and that MT1-MMP is a critical mediator of tendon fibroblast migration and a potential target for the treatment of tendon injuries and diseases.

scholarly journals Reciprocal regulation among TRPV1 channels and phosphoinositide 3-kinase in response to nerve growth factor

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Anastasiia Stratiievska ◽  
Sara Nelson ◽  
Eric N Senning ◽  
Jonathan D Lautz ◽  
Stephen EP Smith ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Total Internal Reflection ◽  
Inflammatory Mediator ◽  
Pi3k Signaling ◽  
Reciprocal Regulation ◽  
Trpv1 Channels ◽  
Pi3k Activity ◽  
Trpv Channels ◽  
Receptor Neurons ◽  
Phosphoinositide 3 Kinase

Although it has been known for over a decade that the inflammatory mediator NGF sensitizes pain-receptor neurons through increased trafficking of TRPV1 channels to the plasma membrane, the mechanism by which this occurs remains mysterious. NGF activates phosphoinositide 3-kinase (PI3K), the enzyme that generates PI(3,4)P2 and PIP3, and PI3K activity is required for sensitization. One tantalizing hint came from the finding that the N-terminal region of TRPV1 interacts directly with PI3K. Using two-color total internal reflection fluorescence microscopy, we show that TRPV1 potentiates NGF-induced PI3K activity. A soluble TRPV1 fragment corresponding to the N-terminal Ankyrin repeats domain (ARD) was sufficient to produce this potentiation, indicating that allosteric regulation was involved. Further, other TRPV channels with conserved ARDs also potentiated NGF-induced PI3K activity. Our data demonstrate a novel reciprocal regulation of PI3K signaling by the ARD of TRPV channels.

scholarly journals TTL-Expression Modulates Epithelial Morphogenesis

2021 ◽  
Vol 9 ◽  
Author(s):  
Manuel Müller ◽  
Karina Ringer ◽  
Florian Hub ◽  
Natalia Kamm ◽  
Thomas Worzfeld ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Focal Adhesion ◽  
Cell Shape ◽  
Epithelial Tissue ◽  
Tissue Formation ◽  
Carboxy Terminus ◽  
Migration Patterns ◽  
Epithelial Monolayers ◽  
Tyrosinated Tubulin

Epithelial monolayer formation depends on the architecture and composition of the microtubule cytoskeleton. Microtubules control bidirectional trafficking and determine the positioning of structural cellular proteins. We studied the role of tubulin tyrosination in epithelial cell shape and motility. Tubulin tyrosine ligase (TTL), the enzyme that adds tyrosine to the carboxy terminus of detyrosinated α-tubulin, was depleted or overexpressed in 2D epithelial monolayers as well as in 3D intestinal organoids. We demonstrate qualitatively and quantitatively that in the absence of TTL the cells comprise high levels of detyrosinated tubulin, change their shape into an initial flat morphology and retardedly acquire a differentiated columnar epithelial cell shape. Enhanced adhesion and accelerated migration patterns of TTL-knockout cells combined with reverse effects in TTL-overexpressing cells indicate that the loss of TTL affects the organization of cell adhesion foci. Precipitation of detyrosinated tubulin with focal adhesion scaffold components coincides with increased quantities and persistence of focal adhesion plaques. Our results indicate that the equilibrium between microtubules enriched in detyrosinated or tyrosinated tubulin modulates epithelial tissue formation, cell morphology, and adhesion.

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