Chemoattractant-Stimulated Rac Activation in Wild-Type and Rac2-Deficient Murine Neutrophils: Preferential Activation of Rac2 and Rac2 Gene Dosage Effect on Neutrophil Functions

The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2(-/-) mice have multiple defects, including chemoattractant-stimulated NADPH oxidase activity and chemotaxis, which may result from an overall reduction in cellular R...

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Published inThe Journal of immunology (1950) Vol. 169; no. 9; pp. 5043 - 5051
Main Authors Li, Shijun, Yamauchi, Akira, Marchal, Christophe C, Molitoris, Jason K, Quilliam, Lawrence A, Dinauer, Mary C
Format Journal Article
LanguageEnglish
Published United States Am Assoc Immnol 01.11.2002
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Abstract The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2(-/-) mice have multiple defects, including chemoattractant-stimulated NADPH oxidase activity and chemotaxis, which may result from an overall reduction in cellular Rac or mechanisms that discriminate Rac1 and Rac2. We show that murine neutrophils have similar amounts of Rac1 and Rac2, unlike human neutrophils, which express predominantly Rac2. An affinity precipitation assay for Rac-GTP showed that although FMLP-induced activation of both isoforms in wild-type neutrophils, approximately 4-fold more Rac2-GTP was detected than Rac1-GTP. Wild-type and Rac2-deficient neutrophils have similar levels of total Rac1. FMLP-induced Rac1-GTP in rac2(-/-) neutrophils was approximately 3-fold greater than in wild-type cells, which have similar levels of total Rac1, yet FMLP-stimulated F-actin, chemotaxis, and superoxide production are markedly impaired in rac2(-/-) neutrophils. Heterozygous rac2(+/-) neutrophils, which had intermediate levels of total and FMLP-induced activated Rac2, exhibited intermediate functional responses to FMLP, suggesting that Rac2 was rate limiting for these functions. Thus, phenotypic defects in FMLP-stimulated Rac2-deficient neutrophils appear to reflect distinct activation and signaling profiles of Rac1 and Rac2, rather than a reduction in the total cellular level of Rac.
AbstractList The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2(-/-) mice have multiple defects, including chemoattractant-stimulated NADPH oxidase activity and chemotaxis, which may result from an overall reduction in cellular Rac or mechanisms that discriminate Rac1 and Rac2. We show that murine neutrophils have similar amounts of Rac1 and Rac2, unlike human neutrophils, which express predominantly Rac2. An affinity precipitation assay for Rac-GTP showed that although FMLP-induced activation of both isoforms in wild-type neutrophils, approximately 4-fold more Rac2-GTP was detected than Rac1-GTP. Wild-type and Rac2-deficient neutrophils have similar levels of total Rac1. FMLP-induced Rac1-GTP in rac2(-/-) neutrophils was approximately 3-fold greater than in wild-type cells, which have similar levels of total Rac1, yet FMLP-stimulated F-actin, chemotaxis, and superoxide production are markedly impaired in rac2(-/-) neutrophils. Heterozygous rac2(+/-) neutrophils, which had intermediate levels of total and FMLP-induced activated Rac2, exhibited intermediate functional responses to FMLP, suggesting that Rac2 was rate limiting for these functions. Thus, phenotypic defects in FMLP-stimulated Rac2-deficient neutrophils appear to reflect distinct activation and signaling profiles of Rac1 and Rac2, rather than a reduction in the total cellular level of Rac.
The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2−/− mice have multiple defects, including chemoattractant-stimulated NADPH oxidase activity and chemotaxis, which may result from an overall reduction in cellular Rac or mechanisms that discriminate Rac1 and Rac2. We show that murine neutrophils have similar amounts of Rac1 and Rac2, unlike human neutrophils, which express predominantly Rac2. An affinity precipitation assay for Rac-GTP showed that although FMLP-induced activation of both isoforms in wild-type neutrophils, ≈4-fold more Rac2-GTP was detected than Rac1-GTP. Wild-type and Rac2-deficient neutrophils have similar levels of total Rac1. FMLP-induced Rac1-GTP in rac2−/− neutrophils was ≈3-fold greater than in wild-type cells, which have similar levels of total Rac1, yet FMLP-stimulated F-actin, chemotaxis, and superoxide production are markedly impaired in rac2−/− neutrophils. Heterozygous rac2+/− neutrophils, which had intermediate levels of total and FMLP-induced activated Rac2, exhibited intermediate functional responses to FMLP, suggesting that Rac2 was rate limiting for these functions. Thus, phenotypic defects in FMLP-stimulated Rac2-deficient neutrophils appear to reflect distinct activation and signaling profiles of Rac 1 and Rac2, rather than a reduction in the total cellular level of Rac.
The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2 super(-/-) mice have multiple defects, including chemoattractant-stimulated NADPH oxidase activity and chemotaxis, which may result from an overall reduction in cellular Rac or mechanisms that discriminate Rac1 and Rac2. We show that murine neutrophils have similar amounts of Rac1 and Rac2, unlike human neutrophils, which express predominantly Rac2. An affinity precipitation assay for Rac-GTP showed that although FMLP-induced activation of both isoforms in wild-type neutrophils, [approx]4-fold more Rac2-GTP was detected than Rac1-GTP. Wild-type and Rac2-deficient neutrophils have similar levels of total Rac1. FMLP-induced Rac1-GTP in rac2 super(-/-) neutrophils was [approx]3-fold greater than in wild-type cells, which have similar levels of total Rac1, yet FMLP-stimulated F-actin, chemotaxis, and superoxide production are markedly impaired in rac2 super(-/-) neutrophils. Heterozygous rac2 super(+/-) neutrophils, which had intermediate levels of total and FMLP-induced activated Rac2, exhibited intermediate functional responses to FMLP, suggesting that Rac2 was rate limiting for these functions. Thus, phenotypic defects in FMLP-stimulated Rac2-deficient neutrophils appear to reflect distinct activation and signaling profiles of Rac 1 and Rac2, rather than a reduction in the total cellular level of Rac.
Author Quilliam, Lawrence A
Marchal, Christophe C
Molitoris, Jason K
Yamauchi, Akira
Dinauer, Mary C
Li, Shijun
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Snippet The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2(-/-) mice have multiple...
The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2−/− mice have multiple...
The hemopoietic-specific Rho family GTPase Rac2 shares 92% amino acid identity with ubiquitously expressed Rac1. Neutrophils from rac2 super(-/-) mice have...
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SubjectTerms Actins - metabolism
Animals
Chemotaxis, Leukocyte - genetics
Chemotaxis, Leukocyte - immunology
Enzyme Inhibitors - pharmacology
Female
Gene Dosage
Genetic Carrier Screening
Humans
Male
Mice
Mice, Inbred C57BL
Mice, Knockout
N-Formylmethionine Leucyl-Phenylalanine - pharmacology
NADPH Oxidases - deficiency
NADPH Oxidases - genetics
Neutrophil Activation - genetics
Neutrophil Activation - immunology
Neutrophils - enzymology
Neutrophils - immunology
Neutrophils - metabolism
Phosphoinositide-3 Kinase Inhibitors
Protein Isoforms - biosynthesis
Protein Isoforms - deficiency
Protein Isoforms - genetics
Protein Isoforms - metabolism
rac GTP-Binding Proteins - biosynthesis
rac GTP-Binding Proteins - deficiency
rac GTP-Binding Proteins - genetics
rac GTP-Binding Proteins - metabolism
rac1 GTP-Binding Protein - biosynthesis
rac1 GTP-Binding Protein - metabolism
RAC2 GTP-Binding Protein
src-Family Kinases - antagonists & inhibitors
Title Chemoattractant-Stimulated Rac Activation in Wild-Type and Rac2-Deficient Murine Neutrophils: Preferential Activation of Rac2 and Rac2 Gene Dosage Effect on Neutrophil Functions
URI http://www.jimmunol.org/cgi/content/abstract/169/9/5043
https://www.ncbi.nlm.nih.gov/pubmed/12391220
https://search.proquest.com/docview/18501558
https://search.proquest.com/docview/72193841
Volume 169
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