title: Retrograde flow and myosin II activity within the leading cell edge deliver F-actin to the lamella to seed the formation of graded polarity actomyosin II filament bundles in migrating fibroblasts.
abstract: In migrating fibroblasts actomyosin II bundles are graded polarity (GP) bundles, a distinct organization to stress fibers. GP bundles are important for powering cell migration, yet have an unknown mechanism of formation. Electron microscopy and the fate of photobleached marks show actin filaments undergoing retrograde flow in filopodia, and the lamellipodium are structurally and dynamically linked with stationary GP bundles within the lamella. An individual filopodium initially protrudes, but then becomes separated from the tip of the lamellipodium and seeds the formation of a new GP bundle within the lamella. In individual live cells expressing both GFP-myosin II and RFP-actin, myosin II puncta localize to the base of an individual filopodium an average 28 s before the filopodium seeds the formation of a new GP bundle. Associated myosin II is stationary with respect to the substratum in new GP bundles. Inhibition of myosin II motor activity in live cells blocks appearance of new GP bundles in the lamella, without inhibition of cell protrusion in the same timescale. We conclude retrograde F-actin flow and myosin II activity within the leading cell edge delivers F-actin to the lamella to seed the formation of new GP bundles.
authors.affiliation: MRC-Laboratory Molecular Cell Biology, University College London, London WC1E 6BT, United Kingdom.
authors.forename: Tom W
authors.initials: TW
authors.lastname: Anderson
authors.forename: Andrew N
authors.initials: AN
authors.lastname: Vaughan
authors.forename: Louise P
authors.initials: LP
authors.lastname: Cramer
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D000199:Actins / Q000378:metabolism*; D000205:Actomyosin / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002465:Cell Movement; D016764:Cell Polarity; D002470:Cell Survival; D002642:Chick Embryo; D005347:Fibroblasts / Q000166:cytology* / Q000648:ultrastructure; D008954:Models, Biological; D024462:Myosin Type II / Q000378:metabolism*; D021122:Protein Subunits / Q000378:metabolism; D011554:Pseudopodia / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D000199:Actins; D021122:Protein Subunits; D000205:Actomyosin; D024462:Myosin Type II
references.citation: J Cell Biol. 1997 Oct 20;139(2):397-415
references.pmid: 9334344
references.citation: Methods Cell Biol. 2003;71:387-416
references.pmid: 12884701
references.citation: Curr Opin Cell Biol. 2006 Feb;18(1):18-25
references.pmid: 16337369
references.citation: J Cell Sci. 2006 Oct 15;119(Pt 20):4342-52
references.pmid: 17038547
references.citation: J Cell Biol. 2003 Feb 3;160(3):409-21
references.pmid: 12566431
references.citation: J Cell Biol. 1992 Dec;119(5):1219-43
references.pmid: 1447299
references.citation: J Cell Sci. 1997 Jan;110 ( Pt 2):123-37
references.pmid: 9044043
references.citation: J Cell Biol. 1995 Nov;131(4):989-1002
references.pmid: 7490299
references.citation: Science. 2003 Dec 5;302(5651):1704-9
references.pmid: 14657486
references.citation: J Cell Biol. 2008 Mar 24;180(6):1233-44
references.pmid: 18362182
references.citation: J Muscle Res Cell Motil. 1980 Mar;1(1):5-14
references.pmid: 7014630
references.citation: J Cell Biol. 2006 May 8;173(3):383-94
references.pmid: 16651381
references.citation: J Cell Biol. 1996 Jun;133(6):1403-15
references.pmid: 8682874
references.citation: Biochem Soc Symp. 1999;65:233-43
references.pmid: 10320942
references.citation: Science. 2003 Mar 14;299(5613):1743-7
references.pmid: 12637748
references.citation: Mol Biol Cell. 2008 Jan;19(1):318-26
references.pmid: 17959823
references.citation: J Cell Sci. 1979 Oct;39:149-65
references.pmid: 575139
references.citation: Cell Motil Cytoskeleton. 2004 Jun;58(2):96-103
references.pmid: 15083531
references.citation: Curr Biol. 2005 Apr 26;15(8):R310-1
references.pmid: 15854900
references.citation: Am J Physiol Renal Physiol. 2005 Sep;289(3):F638-43
references.pmid: 15886277
references.citation: J Cell Biol. 1999 Sep 6;146(5):1097-106
references.pmid: 10477762
references.citation: Sci STKE. 2007 Aug 21;2007(400):re5
references.pmid: 17712139
references.citation: Neuron. 1995 Apr;14(4):763-71
references.pmid: 7536426
references.citation: Cell. 2003 Feb 21;112(4):453-65
references.pmid: 12600310
references.citation: Cell. 1996 Feb 9;84(3):371-9
references.pmid: 8608590
references.citation: Front Biosci. 1997 Jun 01;2:d260-70
references.pmid: 9206973
references.citation: J Cell Biol. 2002 Jul 8;158(1):139-52
references.pmid: 12105186
references.citation: J Cell Biol. 1997 Aug 25;138(4):783-97
references.pmid: 9265646
references.citation: J Cell Sci. 2007 Oct 15;120(Pt 20):3491-9
references.pmid: 17928305
references.citation: Curr Biol. 1998 Dec 17-31;8(25):1347-56
references.pmid: 9889097
references.citation: J Cell Biol. 1998 Sep 7;142(5):1301-12
references.pmid: 9732290
references.citation: Proc Natl Acad Sci U S A. 2004 Jun 29;101(26):9660-5
references.pmid: 15210979
references.citation: J Cell Sci. 1983 Mar;60:331-54
references.pmid: 6348051
references.citation: Cell Motil Cytoskeleton. 1990;16(1):14-21
references.pmid: 2354525
references.citation: Biochem Soc Symp. 1999;65:207-22
references.pmid: 10320940
references.citation: Biochim Biophys Acta. 1998 Sep 16;1404(3):271-81
references.pmid: 9739149
references.citation: Biochem Soc Symp. 1999;65:173-205
references.pmid: 10320939
references.citation: Cell Motil Cytoskeleton. 2002 Jan;51(1):27-38
references.pmid: 11810694
references.citation: Cell. 1996 Feb 9;84(3):359-69
references.pmid: 8608589
references.citation: Curr Opin Cell Biol. 2005 Oct;17(5):517-23
references.pmid: 16099152
references.citation: J Cell Biol. 2002 Jul 8;158(1):31-7
references.pmid: 12105180
references.citation: J Anat. 2004 Nov;205(5):381-91
references.pmid: 15575887
references.citation: J Cell Biol. 1989 Sep;109(3):1163-72
references.pmid: 2670956
references.citation: Cell Motil Cytoskeleton. 1991;18(4):245-57
references.pmid: 2049790
references.citation: Mol Biol Cell. 1999 Dec;10(12):4075-90
references.pmid: 10588644
references.citation: Cell. 2004 Feb 6;116(3):431-43
references.pmid: 15016377
references.citation: Cell. 2007 Feb 9;128(3):561-75
references.pmid: 17289574
references.citation: J Mol Biol. 2008 Jan 11;375(2):325-30
references.pmid: 18021803
references.citation: J Cell Biol. 1997 Mar 24;136(6):1287-305
references.pmid: 9087444
references.citation: Mol Biol Cell. 2005 Mar;16(3):1223-31
references.pmid: 15635099
references.citation: Trends Cell Biol. 1995 Feb;5(2):52-5
references.pmid: 14731405
references.citation: Mol Biol Cell. 2005 Feb;16(2):902-17
references.pmid: 15591129
references.citation: Semin Cell Biol. 1994 Jun;5(3):149-55
references.pmid: 7919228
references.citation: Curr Biol. 2003 Feb 4;13(3):252-7
references.pmid: 12573223
references.citation: J Cell Biol. 1988 Dec;107(6 Pt 2):2631-45
references.pmid: 3204122
references.citation: Mol Biol Cell. 2007 Oct;18(10):3723-32
references.pmid: 17634292
references.citation: J Biol Chem. 2007 Aug 10;282(32):23005-9
references.pmid: 17576764
references.citation: Neuron. 1996 Apr;16(4):769-82
references.pmid: 8607995
references.citation: Curr Biol. 2002 Aug 20;12(16):1413-18
references.pmid: 12194823
references.citation: Nat Cell Biol. 2006 Mar;8(3):215-26
references.pmid: 16501565
references.citation: Symp Soc Exp Biol. 1993;47:35-56
references.pmid: 8165576
references.citation: Curr Biol. 1999 Oct 7;9(19):1095-105
references.pmid: 10531004
references.citation: Nat Rev Mol Cell Biol. 2008 Jun;9(6):446-54
references.pmid: 18464790
references.citation: J Cell Biol. 1993 Mar;120(6):1381-91
references.pmid: 8449984