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 cult ured epithelia, the proper organization of the actin filament ring, te thered to the plasma membrane at the zonula adhaerens, is apparently n ecessary for their functioning as a transporting epithelium. It has be en 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 prote ins required for cell polarization. To test this hypothesis, the autho rs analyzed the localization and possible association of the actin-bin ding motor protein myosin I with actin filaments during changes in the actin ring position and organization, and also with trans-Golgi-deriv ed vesicles. Modifications of the ring were induced subjecting the cel ls to external Ca2+ depletion and restoration (Ca2+ switch), or by tre atment with drugs known to depolymerize actin filaments (cytochalasin D, CD). The distribution of myosin I and actin, both in intact cells a nd in cellular fractions, was monitored using heterologous crossreacti ng antibodies and phalloidin. The authors identified an isoform of myo sin I of approximately 110-125 KDa, homologous to myosin IB of Acantha moeba, 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 co localized,vith the actin fibers but appeared dispersed in the cytoplas m. Furthermore, a significant fraction of the total myosin I in the ce ll was associated to Golgi-derived vesicles which could also associate in vitro with actin filaments. The authors' data support, then, the p articipation of myosin I, in association with actin filaments, in vesi cle translocation to and from the cell membrane as proposed for natura l epithelia, and provide a further insight into the structural organiz ation that maintains epithelial cell polarity in cultured monolayers.