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p21-Activated kinase (PAK) regulates cytoskeletal reorganization and directional migration in human neutrophils.

Itakura A, Aslan JE, Kusanto BT, Phillips KG, Porter JE, Newton PK, Nan X, Insall RH, Chernoff J, McCarty OJ - PLoS ONE (2013)

Bottom Line: In this study, we characterized the role of p21-activated kinase (PAK) downstream of Rho GTPases in cytoskeletal remodeling and chemotactic processes of human neutrophils.We found that PAK activation occurred upon stimulation of neutrophils with f-Met-Leu-Phe (fMLP), and PAK accumulated at the actin-rich leading edge of stimulated neutrophils, suggesting a role for PAK in Rac-dependent actin remodeling.Treatment with the pharmacological PAK inhibitor, PF3758309, abrogated the integrity of RhoA-mediated actomyosin contractility and surface adhesion.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Oregon Health and Science University, Portland, Oregon, United States of America.

ABSTRACT
Neutrophils serve as a first line of defense in innate immunity owing in part to their ability to rapidly migrate towards chemotactic factors derived from invading pathogens. As a migratory function, neutrophil chemotaxis is regulated by the Rho family of small GTPases. However, the mechanisms by which Rho GTPases orchestrate cytoskeletal dynamics in migrating neutrophils remain ill-defined. In this study, we characterized the role of p21-activated kinase (PAK) downstream of Rho GTPases in cytoskeletal remodeling and chemotactic processes of human neutrophils. We found that PAK activation occurred upon stimulation of neutrophils with f-Met-Leu-Phe (fMLP), and PAK accumulated at the actin-rich leading edge of stimulated neutrophils, suggesting a role for PAK in Rac-dependent actin remodeling. Treatment with the pharmacological PAK inhibitor, PF3758309, abrogated the integrity of RhoA-mediated actomyosin contractility and surface adhesion. Moreover, inhibition of PAK activity impaired neutrophil morphological polarization and directional migration under a gradient of fMLP, and was associated with dysregulated Ca(2+) signaling. These results suggest that PAK serves as an important effector of Rho-family GTPases in neutrophil cytoskeletal reorganization, and plays a key role in driving efficient directional migration of human neutrophils.

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A model for PAK-mediated cytoskeletal regulation in human neutrophils.PAK coordinates the crosstalk between Rho GTPases and cytoskeletal dynamics in migrating neutrophils.
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pone-0073063-g006: A model for PAK-mediated cytoskeletal regulation in human neutrophils.PAK coordinates the crosstalk between Rho GTPases and cytoskeletal dynamics in migrating neutrophils.

Mentions: The efficient migratory ability of neutrophils requires a network of interactions between distinct cytoskeletal signaling systems, namely frontness signals regulated by actin-dependent membrane protrusion and backness signals mediated by myosin-dependent cell contraction. Recent studies suggest that crosstalk processes between frontness and backness modules during the initiation, establishment and maintenance of neutrophil polarization are dynamic and complex [41]. Moreover, the mediators of temporal and spatial regulation in this crosstalk network remain to be elucidated. In this study, we identified the p21-activated kinase PAK as a regulator of neutrophil cytoskeletal reorganization. The PAK family proteins serve as effectors downstream of Rac and Cdc42 GTPases and are noted for roles in cytoskeletal dynamics, gene transcription, survival signaling, and cell cycle progression [25], [23]. Here we find that the kinase activity of PAK is required for the efficient directional migration of human neutrophils, where PAK2 may serve key roles in frontness signals at the leading edge. The absence of PAK activity resulted in an impairment of polarization, which was associated with increased backness signals and surface adhesions. Together, as depicted in Fig. 6, our data describe a differential spatial regulation of PAK isoforms in fMLP-stimulated human neutrophils and highlights a potential mechanism for PAK-mediated crosstalk between Rho GTPases during cytoskeletal reorganization.


p21-Activated kinase (PAK) regulates cytoskeletal reorganization and directional migration in human neutrophils.

Itakura A, Aslan JE, Kusanto BT, Phillips KG, Porter JE, Newton PK, Nan X, Insall RH, Chernoff J, McCarty OJ - PLoS ONE (2013)

A model for PAK-mediated cytoskeletal regulation in human neutrophils.PAK coordinates the crosstalk between Rho GTPases and cytoskeletal dynamics in migrating neutrophils.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3760889&req=5

pone-0073063-g006: A model for PAK-mediated cytoskeletal regulation in human neutrophils.PAK coordinates the crosstalk between Rho GTPases and cytoskeletal dynamics in migrating neutrophils.
Mentions: The efficient migratory ability of neutrophils requires a network of interactions between distinct cytoskeletal signaling systems, namely frontness signals regulated by actin-dependent membrane protrusion and backness signals mediated by myosin-dependent cell contraction. Recent studies suggest that crosstalk processes between frontness and backness modules during the initiation, establishment and maintenance of neutrophil polarization are dynamic and complex [41]. Moreover, the mediators of temporal and spatial regulation in this crosstalk network remain to be elucidated. In this study, we identified the p21-activated kinase PAK as a regulator of neutrophil cytoskeletal reorganization. The PAK family proteins serve as effectors downstream of Rac and Cdc42 GTPases and are noted for roles in cytoskeletal dynamics, gene transcription, survival signaling, and cell cycle progression [25], [23]. Here we find that the kinase activity of PAK is required for the efficient directional migration of human neutrophils, where PAK2 may serve key roles in frontness signals at the leading edge. The absence of PAK activity resulted in an impairment of polarization, which was associated with increased backness signals and surface adhesions. Together, as depicted in Fig. 6, our data describe a differential spatial regulation of PAK isoforms in fMLP-stimulated human neutrophils and highlights a potential mechanism for PAK-mediated crosstalk between Rho GTPases during cytoskeletal reorganization.

Bottom Line: In this study, we characterized the role of p21-activated kinase (PAK) downstream of Rho GTPases in cytoskeletal remodeling and chemotactic processes of human neutrophils.We found that PAK activation occurred upon stimulation of neutrophils with f-Met-Leu-Phe (fMLP), and PAK accumulated at the actin-rich leading edge of stimulated neutrophils, suggesting a role for PAK in Rac-dependent actin remodeling.Treatment with the pharmacological PAK inhibitor, PF3758309, abrogated the integrity of RhoA-mediated actomyosin contractility and surface adhesion.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Oregon Health and Science University, Portland, Oregon, United States of America.

ABSTRACT
Neutrophils serve as a first line of defense in innate immunity owing in part to their ability to rapidly migrate towards chemotactic factors derived from invading pathogens. As a migratory function, neutrophil chemotaxis is regulated by the Rho family of small GTPases. However, the mechanisms by which Rho GTPases orchestrate cytoskeletal dynamics in migrating neutrophils remain ill-defined. In this study, we characterized the role of p21-activated kinase (PAK) downstream of Rho GTPases in cytoskeletal remodeling and chemotactic processes of human neutrophils. We found that PAK activation occurred upon stimulation of neutrophils with f-Met-Leu-Phe (fMLP), and PAK accumulated at the actin-rich leading edge of stimulated neutrophils, suggesting a role for PAK in Rac-dependent actin remodeling. Treatment with the pharmacological PAK inhibitor, PF3758309, abrogated the integrity of RhoA-mediated actomyosin contractility and surface adhesion. Moreover, inhibition of PAK activity impaired neutrophil morphological polarization and directional migration under a gradient of fMLP, and was associated with dysregulated Ca(2+) signaling. These results suggest that PAK serves as an important effector of Rho-family GTPases in neutrophil cytoskeletal reorganization, and plays a key role in driving efficient directional migration of human neutrophils.

Show MeSH
Related in: MedlinePlus