Actin remodeling in motile cells

Non-muscle cell shape change and motility depend primarily on the dynamics and distributions of cytoplasmic actin. In cells, actin cycles between monomeric and polymeric phases tightly regulated by actin binding proteins that control cellular architecture and movement. Here, we characterize actin remodeling in shear stress stimulated endothelial cells and in actin networks reconstituted with purified proteins. Fluid shear stress stimulation induces endothelial cells to elongate and align in the direction of applied flow. Alignment requires 24 h of exposure to flow, but the cells respond within minutes to flow by diminishing their movements by 50%. Although movement slows, actin filament turnover times and the amount of polymerized actin in cells decreases, increasing actin filament remodeling in individual cells composing a confluent endothelial monolayer to levels used by disperse, non-confluent cells for rapid movement. Hours later, motility returns to pre-shear stress levels, but actin remodeling remains highly dynamic in many cells. We conclude that shear stress initiates a cytoplasmic actin remodeling response that is used to modify endothelial cell shape instead of bulk cell translocation. We determine the steady state dynamics of purified actin filament networks in the entangled state and after orthogonal cross-linking with filamins using a novel, non-perturbing fluorescence system. Human filamin A or Dictyosteliun discoidium filamin slow actin filament turnover by [approximately] 50% and recruit much of a significant population of actin oligomers that we measure are present in polymerized purified actin solutions into the immobile filament fraction. Surprisingly, these observations occur at very low stoichiometry to actin, approximately requiring only one

Department dc:contributor.department

Harvard University--MIT Division of Health Sciences and Technology.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2004

Author dc:creator

• Osborn, Eric A. (Eric Alan), 1975-

Advisor dc:contributor.advisor

• C. Forbes Dewey.

• Harvard University--MIT Division of Health Sciences and Technology.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

en_US

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/28600

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/28600