Differential binding to dorsal and ventral cell surfaces of fibroblasts: effect on collagen phagocytosis.
Matrix remodeling by phagocytic fibroblasts is essential for growth and development but the regulatory processes are undefined. We evaluated the impact of spreading on the binding step of collagen phagocytosis with a novel culture system that more closely replicates phagocytosis in vivo than previous models. 3T3 cells were plated on collagen-coated beads, thereby loading only ventral surfaces (adhesion with spreading), or were allowed to spread on collagen films and then loaded with beads on their dorsal surfaces (adhesion without spreading). Ventral surfaces bound three-fold more beads than dorsal surfaces which was accompanied by accelerated phagosomal maturation. Arp3 and cortactin, markers of the actin-associated spreading machinery, strongly accumulated around ventrally but not dorsally loaded beads, suggesting that spreading contributes to enhanced binding of ventral surfaces. Further, ventral surfaces exhibited two-fold more free alpha2beta1 integrins, the major collagen receptors. Notably, compared to cells spread on collagen substrates, spreading cells exhibited a three-fold higher alpha2beta1 mobile fraction which was correlated with limited engagement of ventral receptors by actin filaments. Thus integrin ligation by actin filaments regulates the mobility of collagen receptors which in turn mediates the enhanced binding of collagen beads on spreading surfaces.
CIHR Group in Matrix Dynamics, Faculty of Dentistry, University of Toronto, Toronto, Ontario M5S 3EZ, Canada.
Pamela D
PD
Arora
Lingzhi
L
Fan
Jaro
J
Sodek
András
A
Kapus
Christopher A
CA
McCulloch
D016475:3T3 Cells; D008841:Actin Cytoskeleton / Q000378:metabolism; D051378:Actin-Related Protein 3; D000818:Animals; D001665:Binding Sites / Q000187:drug effects / Q000502:physiology; D002448:Cell Adhesion / Q000187:drug effects* / Q000502:physiology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects* / Q000502:physiology; D016764:Cell Polarity / Q000187:drug effects / Q000502:physiology; D003094:Collagen / Q000378:metabolism / Q000494:pharmacology*; D046508:Culture Techniques / Q000379:methods*; D003598:Cytoskeletal Proteins / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D038982:Integrin alpha2beta1 / Q000378:metabolism; D051379:Mice; D008855:Microscopy, Electron, Scanning; D008863:Microspheres; D010587:Phagocytosis / Q000187:drug effects* / Q000502:physiology; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology
D051378:Actin-Related Protein 3; C495929:Actr3 protein, mouse; D003598:Cytoskeletal Proteins; D038982:Integrin alpha2beta1; D003094:Collagen