Myosin I interactions with actin filaments and trans-Golgi-derived vesicles in MDCK cell monolayers.
In MDCK cell cultured monolayers, as well as in natural and other cultured epithelia, the proper organization of the actin filament ring, tethered to the plasma membrane at the zonula adhaerens, is apparently necessary for their functioning as a transporting epithelium. It has been proposed that actin filaments, in conjunction with motor proteins, could provide the structural basis that regulates the tight junction (TJ) sealing capacity as well as the transport of membrane-tagged proteins required for cell polarization. To test this hypothesis, the authors analyzed the localization and possible association of the actin-binding motor protein myosin I with actin filaments during changes in the actin ring position and organization, and also with trans-Golgi-derived vesicles. Modifications of the ring were induced subjecting the cells to external Ca2+ depletion and restoration (Ca2+ switch), or by treatment with drugs known to depolymerize actin filaments (cytochalasin D, CD). The distribution of myosin I and actin, both in intact cells and in cellular fractions, was monitored using heterologous cross-reacting antibodies and phalloidin. The authors identified an isoform of myosin I of approximately 110-125 KDa, homologous to myosin IB of Acanthamoeba, a fraction of which colocalized with the peripheral actin ring. The association seems transient as, once the ring retracted as result of Ca2+ depletion, or became disorganized by CD, myosin not longer colocalized with the actin fibers but appeared dispersed in the cytoplasm. Furthermore, a significant fraction of the total myosin I in the cell was associated to Golgi-derived vesicles which could also associate in vitro with actin filaments. The authors' data support, then, the participation of myosin I, in association with actin filaments, in vesicle translocation to and from the cell membrane as proposed for natural epithelia, and provide a further insight into the structural organization that maintains epithelial cell polarity in cultured monolayers.
Montes de Oca
G
G
Departamento de Biología Celular y Programa de Biomedicina Molecular, Centro de Investigación y de Estudios Avanzados, (CINVESTAV) del Instituto Politécnico Nacional, México, D.F.
Lezama
R A
RA
Mondragón
R
R
Castillo
A M
AM
Meza
I
I
D008841:Actin Cytoskeleton / Q000378:metabolism*; D000199:Actins / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D004285:Dogs; D006056:Golgi Apparatus / Q000378:metabolism*; D007686:Kidney Tubules, Distal / Q000378:metabolism / Q000648:ultrastructure; D009218:Myosins / Q000378:metabolism*
D000199:Actins; D009218:Myosins