TY - JOUR
T1 - Characterization and localization of myosin in the brush border of intestinal epithelial cells
AU - Mooseker, Mark S.
AU - Pollard, Thomas D.
AU - Fujiwara, Keigi
PY - 1978/11/1
Y1 - 1978/11/1
N2 - The brush border of intestinal epithelial cells consists of a tightly packed array of microvilli, each of which contains a core of actin filaments. It has been postulated that microvillar movements are mediated by myosin interactions in the terminal web with the basal ends of these actin cores (Mooseker, M. S. 1976. /. Cell Biol. 71:417-433). We report here that two predictions of this model are correct: (a) The brush border contains myosin, and (b) myosin is located in the terminal web. Myosin is isolated in 70% purity by solubilization of Triton-treated brush borders in 0.6 M KI, and separation of the components by gel filtration. Most of the remaining contaminants can be removed by precipitation of the myosin at low ionic strength. The yield is ~1 mg of myosin/30 mg of solubilized brush border protein. The molecule consists of three subunits with molecular weights of 200,000, 19,000, and 17,000 daltons in a 1:1:1 M ratio. At low ionic strength, the myosin forms small, bipolar filaments with dimensions of 300 × 11 nm, that are similar to filaments seen previously in the terminal web of isolated brush borders. Like that of other vertebrate, nonmuscle myosins, the ATPase activity of isolated brush border myosin in 0.6 M KC1 is highest with EDTA (1 µmol P1/ mg-min; 37°C), intermediate with Ca++ (0.4 µmol P,/mg-min), and low with Mg++ (0.01 µmol E/mg-min). Actin does not stimulate the Mg-ATPase activity of the isolated enzyme. Antibodies against the rod fragment of human platelet myosin cross-react by immunodiffusion with brush border myosin. Staining of isolated mouse or chicken brush borders with rhodamine-antimyosin demonstrates that myosin is localized exclusively in the terminal web.
AB - The brush border of intestinal epithelial cells consists of a tightly packed array of microvilli, each of which contains a core of actin filaments. It has been postulated that microvillar movements are mediated by myosin interactions in the terminal web with the basal ends of these actin cores (Mooseker, M. S. 1976. /. Cell Biol. 71:417-433). We report here that two predictions of this model are correct: (a) The brush border contains myosin, and (b) myosin is located in the terminal web. Myosin is isolated in 70% purity by solubilization of Triton-treated brush borders in 0.6 M KI, and separation of the components by gel filtration. Most of the remaining contaminants can be removed by precipitation of the myosin at low ionic strength. The yield is ~1 mg of myosin/30 mg of solubilized brush border protein. The molecule consists of three subunits with molecular weights of 200,000, 19,000, and 17,000 daltons in a 1:1:1 M ratio. At low ionic strength, the myosin forms small, bipolar filaments with dimensions of 300 × 11 nm, that are similar to filaments seen previously in the terminal web of isolated brush borders. Like that of other vertebrate, nonmuscle myosins, the ATPase activity of isolated brush border myosin in 0.6 M KC1 is highest with EDTA (1 µmol P1/ mg-min; 37°C), intermediate with Ca++ (0.4 µmol P,/mg-min), and low with Mg++ (0.01 µmol E/mg-min). Actin does not stimulate the Mg-ATPase activity of the isolated enzyme. Antibodies against the rod fragment of human platelet myosin cross-react by immunodiffusion with brush border myosin. Staining of isolated mouse or chicken brush borders with rhodamine-antimyosin demonstrates that myosin is localized exclusively in the terminal web.
KW - Antibody ·
KW - Brush border ·
KW - Intestine ·
KW - Myosin ·
KW - Terminal web
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U2 - 10.1083/jcb.79.2.444
DO - 10.1083/jcb.79.2.444
M3 - Article
C2 - 152766
AN - SCOPUS:0018094857
SN - 0021-9525
VL - 79
SP - 444
EP - 453
JO - Journal of Cell Biology
JF - Journal of Cell Biology
IS - 2
ER -