Primary granule exocytosis in human neutrophils is regulated by Rac-dependent actin remodeling.
The actin cytoskeleton regulates exocytosis in all secretory cells. In neutrophils, Rac2 GTPase has been shown to control primary (azurophilic) granule exocytosis. In this report, we propose that Rac2 is required for actin cytoskeletal remodeling to promote primary granule exocytosis. Treatment of neutrophils with low doses (< or = 10 microM) of the actin-depolymerizing drugs latrunculin B (Lat B) or cytochalasin B (CB) enhanced both formyl peptide receptor- and Ca(2+) ionophore-stimulated exocytosis. Higher concentrations of CB or Lat B, or stabilization of F-actin with jasplakinolide (JP), inhibited primary granule exocytosis measured as myeloperoxidase release but did not affect secondary granule exocytosis determined by lactoferrin release. These results suggest an obligatory role for F-actin disassembly before primary granule exocytosis. However, lysates from secretagogue-stimulated neutrophils showed enhanced actin polymerization activity in vitro. Microscopic analysis showed that resting neutrophils contain significant cortical F-actin, which was redistributed to sites of primary granule translocation when stimulated. Exocytosis and actin remodeling was highly polarized when cells were primed with CB; however, polarization was reduced by Lat B preincubation, and both polarization and exocytosis were blocked when F-actin was stabilized with JP. Treatment of cells with the small molecule Rac inhibitor NSC23766 also inhibited actin remodeling and primary granule exocytosis induced by Lat B/fMLF or CB/fMLF, but not by Ca(2+) ionophore. Therefore, we propose a role for F-actin depolymerization at the cell cortex coupled with Rac-dependent F-actin polymerization in the cell cytoplasm to promote primary granule exocytosis.
Mitchell
Troy
T
Dept. of Cell Biology, Univ. of Alberta, Edmonton, Alberta, Canada T6G 2H7.
Lo
Andrea
A
Logan
Michael R
MR
Lacy
Paige
P
Eitzen
Gary
G
D000199:Actins / Q000378:metabolism*; D000634:Aminoquinolines / Q000494:pharmacology; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D003571:Cytochalasin B / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D047630:Depsipeptides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005089:Exocytosis / Q000187:drug effects; D006801:Humans; D007476:Ionophores / Q000494:pharmacology; D046529:Microscopy, Electron, Transmission; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D011743:Pyrimidines / Q000494:pharmacology; D016897:Respiratory Burst; D022142:Secretory Vesicles / Q000187:drug effects / Q000378:metabolism*; D053778:Thiazolidines / Q000494:pharmacology; D013997:Time Factors; D020744:rac GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000097766:RAC2 GTP-Binding Protein
D000199:Actins; D000634:Aminoquinolines; D019086:Bridged Bicyclo Compounds, Heterocyclic; D000001:Calcimycin; D002118:Calcium; D003571:Cytochalasin B; D047630:Depsipeptides; D004791:Enzyme Inhibitors; D007476:Ionophores; D009240:N-Formylmethionine Leucyl-Phenylalanine; D011743:Pyrimidines; D053778:Thiazolidines; D020744:rac GTP-Binding Proteins; D000097766:RAC2 GTP-Binding Protein; C487513:NSC 23766; C057531:jasplakinolide; C037068:latrunculin B
Blood. 2004 Aug 1;104(3):832-9
15073033
Biochem J. 2007 Mar 1;402(2):229-39
17090228
J Immunol. 2003 Oct 15;171(8):4425-30
14530369
Mol Biol Cell. 1998 May;9(5):1053-63
9571239
Cell. 2003 Jul 25;114(2):201-14
12887922
Curr Top Microbiol Immunol. 2005;291:91-111
15981461
Mol Biol Cell. 2004 Feb;15(2):520-31
14617808
Physiol Rev. 1987 Oct;67(4):1249-95
3317458
Cell Motil Cytoskeleton. 1989;13(3):127-44
2776221
J Cell Biol. 1991 Sep;114(6):1179-90
1716633
Front Biosci. 2008 May 01;13:5559-70
18508605
J Biol Chem. 2007 Oct 19;282(42):30466-75
17726018
J Leukoc Biol. 2003 Mar;73(3):333-8
12629146
J Cell Sci. 1995 Nov;108 ( Pt 11):3547-56
8586666
J Exp Med. 1998 Aug 3;188(3):549-59
9687532
Science. 2003 Oct 17;302(5644):445-9
14564009
Nat Med. 2005 Aug;11(8):886-91
16025125
Nature. 1992 Nov 19;360(6401):270-3
1331813
J Cell Biol. 2003 Nov 10;163(3):559-70
14610058
Proc Natl Acad Sci U S A. 2004 May 18;101(20):7618-23
15128949
J Allergy Clin Immunol. 2003 May;111(5):923-32; quiz 933
12743551
J Leukoc Biol. 1997 Jun;61(6):647-53
9201255
J Immunol. 1978 Sep;121(3):851-5
211165
J Biol Chem. 1994 Dec 9;269(49):30749-52
7982999
Proc Natl Acad Sci U S A. 2001 Jan 30;98(3):1154-9
11158610
J Cell Biol. 1994 Aug;126(4):1005-15
8051203
Immunology. 1999 Oct;98(2):244-52
10540223
J Cell Biol. 2002 Aug 19;158(4):669-79
12177043
Methods Enzymol. 1982;85 Pt B:182-210
6889668
J Cell Biol. 2001 Feb 19;152(4):825-34
11266472
J Clin Invest. 1993 Sep;92(3):1467-76
8376598
Biochim Biophys Acta. 2006 Nov;1763(11):1175-83
17034880
J Biol Chem. 1994 May 27;269(21):14869-71
8195116
Physiol Rev. 2003 Apr;83(2):581-632
12663867
Mol Biol Cell. 1996 Sep;7(9):1429-42
8885237
Am J Physiol Cell Physiol. 2007 May;292(5):C1690-700
17202227
Nat Immunol. 2004 Jul;5(7):744-51
15170212
Curr Biol. 2005 Oct 25;15(20):1867-73
16243035
J Exp Med. 1976 May 1;143(5):1154-69
1262785
J Biol Chem. 1994 Mar 11;269(10):7055-8
8125910
Mol Biol Cell. 2001 Jan;12(1):155-70
11160830
J Biol Chem. 1999 May 7;274(19):13198-204
10224076
Blood. 1999 Aug 1;94(3):1121-30
10419906
J Cell Biol. 1981 Jun;89(3):585-92
7251666
Blood. 2006 Oct 15;108(8):2814-20
16809619
Biophys J. 2000 Jun;78(6):2863-77
10827968
Am J Physiol Cell Physiol. 2000 Sep;279(3):C619-33
10942712
J Cell Biol. 2001 Feb 19;152(4):835-42
11266473
J Cell Biochem. 2000 Sep 18;80(1):37-45
11029752
J Cell Biol. 2000 Aug 21;150(4):785-96
10953003
J Cell Biol. 2007 Oct 22;179(2):239-45
17954607