Microfilament-dependent movement of the beta3 integrin subunit within focal contacts of endothelial cells.
To gain insight into the dynamic properties of focal contacts, we induced expression of green fluorescent protein-tagged beta3 integrin (GFP-beta3) and actinin-1 (GFP-actinin-1) in endothelial cells. Both tagged proteins localize with alpha(v)beta3 integrin in focal contacts distributed towards the periphery of transfected cells. Labeled focal contacts migrate at about 0.1 mm/min in stationary live endothelial cells. We compared beta3 integrin and actinin-1 dynamics in focal contacts by using fluorescence recovery after photobleaching. Recovery of signal in bleached focal contacts that have incorporated actinin-1 is rapid and occurs within less than 4 min. This recovery is energy-dependent. In contrast, recovery of bleached focal contacts that contain GFP-beta3 integrin takes longer than 30 min. Yet, when a narrow stripe of fluorescence is bleached across a beta3 integrin-labeled focal contact, recovery is complete within 16 min. The latter recovery is energy-dependent and is blocked not only by actin-filament disrupting drugs but also by a myosin light chain kinase inhibitor. Thus, integrins are not immobile when incorporated into focal contacts, as some have suggested. We propose that integrins are mobile within the confines of focal contacts and that this mobility is supported by an actin-associated molecular motor.
Tsuruta
Daisuke
D
Department of Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Gonzales
Meredith
M
Hopkinson
Susan B
SB
Otey
Carol
C
Khuon
Satya
S
Goldman
Robert D
RD
Jones
Jonathan C R
JC
D008841:Actin Cytoskeleton / Q000502:physiology*; D000185:Actinin / Q000235:genetics / Q000378:metabolism; D000199:Actins / Q000502:physiology; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology*; D015638:Cytochalasin D / Q000494:pharmacology; D003599:Cytoskeleton / Q000502:physiology; D003847:Deoxyglucose / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D006801:Humans; D039661:Integrin beta3; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D010980:Platelet Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D019810:Sodium Azide / Q000494:pharmacology
D000199:Actins; D015703:Antigens, CD; D039661:Integrin beta3; D008164:Luminescent Proteins; D010980:Platelet Membrane Glycoproteins; D000185:Actinin; D049452:Green Fluorescent Proteins; D015638:Cytochalasin D; D019810:Sodium Azide; D003847:Deoxyglucose