- Cap formation
When
molecules on the surface of a cell arecrosslink ed, they are moved to one end of the cell to form a “cap”. This phenomenon, the process of which is called cap formation, was discovered in 1971 onlymphocytes cite journal | author=Taylor, RB, "et al."| title=Redistribution and pinocytosis of lymphocyte surface immunoglobulin molecules induced by an anti-immunoglobulin antibody| journal=Nature| year=1971| issue=233| pages=225–229] and is a property ofamoebae and all locomotory animal cells except sperm. The crosslinking is most easily achieved using apolyvalent antibody to a surfaceantigen on the cell. Cap formation can be visualised by attaching afluorophore , such asfluorescein , to the antibody.teps in cap formation
#The antibody is bound to the cell. If the antibody is non-crosslinking (such as a Fab antibody fragment), the bound antibody is uniformly distributed. This can be done at 0 °C, room temperature, or 37 °C.
#If the antibody is crosslinking and bound to the cells at 0 °C, the distribution of antibodies has a patchy appearance. These “patches” are two-dimensional precipitates of antigen-antibody complex and are quite analogous to the three-dimensional precipitates which form in solution.
#If cells with patches are warmed up, the patches move to one end of the cell to form a cap. In lymphocytes, this capping process takes about 5 minutes. If carried out on cells attached to a substratum, the cap forms at the rear of the moving cell.Capping only occurs on motile cells and is therefore believed to reflect an intrinsic property of how cells move. It is an energy dependent process and in lymphocytes is partially inhibited by
cytochalasin B (which disruptsmicrofilaments ) but unaffected bycolchicine (which disruptsmicrotubules ). However, a combination of these drugs eliminates capping. A key feature of capping is that only those molecules which are crosslinked cCap formation is now seen as closely related to the carbon particle experiments of Abercrombiecite journal | author=Abercrombie, M, "et al."| title=The locomotion of fibroblasts in culture. III. Movement of particles on the dorsal surface of the leading lamella| journal=Experimental Cell Research| year=1970| volume=62| issue=2-3| url=http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6WFC-4DNHPHP-JV&_user=1861184&_coverDate=10%2F31%2F1970&_alid=502013301&_rdoc=4&_fmt=summary&_orig=search&_cdi=6791&_sort=d&_docanchor=&view=c&_acct=C000055073&_version=1&_urlVersion=0&_userid=1861184&md5=4a5521007d19f99cf01e5e8eec1b85fb/| pages=389–398| doi=10.1016/0014-4827(70)90570-7] . In this case, crawling
fibroblasts were held in a medium containing small (~1 micrometre in size) carbon particles. Occasionally these particles attached to the front leading edge of these cells: when they did so, the particles were observed to move rearwards on the cell’s dorsal surface. They did so in a roughly straight line with the particle remaining initially stationary with respect to the substratum. The cell seemed to ooze forward under the particle. In view of what we know of capping, this phenomenon is now interpreted as follows: the particle is presumably stuck to many surface molecules, crosslinking them and forming a patch. As in capping, the particle moves towards the back of the cell.Proposed mechanisms
"Flow"
Abercrombie thought that the carbon particle is a marker on the cell surface and its behaviour reflects what the surface is doing. This led him to propose that, as a cell moves, membrane from internal stores is added at the front of the cell — enabling the cell to extend forwards — and retrieved towards the rear of the cell. This process of
exocytosis at the front of the cell andendocytosis elsewhere has been modified by Bretschercite journal | author=Bretscher, MS, "et al."| title=Endocytosis: relation to capping and cell locomotion| journal=Experimental Cell Research| year=1984| volume=224| issue=4650| url=http://www.sciencemag.org/cgi/content/abstract/224/4650/681/| pages=681–686| pmid=6719108| doi=10.1126/science.6719108| format=abstract page] cite journal | author=Bretscher, MS| title=Directed lipid flow in cell membranes| journal=Nature| year=1976| volume=260| url=http://www.nature.com/nature/journal/v260/n5546/abs/260021a0.html| pages=21–23| doi=10.1038/260021a0| format=abstract page] cite journal | author=Bretscher, MS| title=Getting Membrane Flow and the Cytoskeleton to Cooperate in Moving Cells| journal=Cell| year=1996| volume=87| issue=4| url=http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6WSN-41SBN1S-4&_coverDate=11%2F15%2F1996&_alid=502022980&_rdoc=1&_fmt=&_orig=search&_qd=1&_cdi=7051&_sort=d&view=c&_acct=C000055073&_version=1&_urlVersion=0&_userid=1861184&md5=3f711c5908778353a328759906e8c333/| pages=601–606| doi=10.1016/S0092-8674(00)81380-X] . He and Hopkinscite journal | author=Hopkins CR, "et al."| title=In migrating fibroblasts, recycling receptors are concentrated in narrow tubules in the pericentriolar area, and then routed to the plasma membrane of the leading lamella| journal=Journal of Cell Biology| year=1994| volume=125| url=http://www.jcb.org/cgi/content/abstract/125/6/1265/| pages=1265–1274| doi=10.1083/jcb.125.6.1265| format=abstract page] showed that the specific membrane endocytosed by coated pits on motile cells is returned byexocytosis to the cell surface at the leading edge. The spatial difference between the sites of exocytosis (at the front) and endocytosis (everywhere on the surface) leads to a flow of the matrix of the plasma membrane — lipids — from the front towards the rear. Large objects, such as patches, would be swept along with this flow whereas non-crosslinked small molecules would be able to diffuse byBrownian motion against the flow and so evade being swept backwards. Hence, in this theory, the need for crosslinking. Bretscher proposed that on stationary cells exocytosis is random — and therefore a major difference between motile and nonmotile cells."Cytoskeleton"
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