Investigation of the role of microtubules in protein secretion from lactating mouse mammary epithelial cells.
Disruption of microtubules has been shown to reduce protein secretion from lactating mammary epithelial cells. To investigate the involvement of microtubules in the secretory pathway in these cells we have examined the effect of nocodazole on protein secretion from mammary epithelial cells derived from the lactating mouse. Mouse mammary cells have extensive microtubule networks and 85% of their tubulin was in a polymeric form. Treatment with 1 micrograms/ml nocodazole converted most of the tubulin into a soluble form. In a continuous labelling protocol it was found that nocodazole did not interfere with protein synthesis but over a 5 h period secretion was markedly inhibited. To determine whether the inhibition was at the level of early or late stages of the secretory pathway mammary cells were pulse-labelled for 1 h to label protein throughout the secretory pathway before nocodazole treatment. When secretion was subsequently assayed it was found to be slower and only partially inhibited. These findings suggest that the major effect of nocodazole is on an early stage of the secretory pathway and that microtubules normally facilitate vesicle transport to the plasma membrane. An involvement of microtubules in vesicle transport to the plasma membrane is consistent with an observed accumulation of casein vesicles in nocodazole-treated cells. Exocytosis stimulated by the calcium ionophore ionomycin was unaffected by nocodazole treatment. We conclude from these results that the major effect of nocodazole is at an early stage of the secretory pathway, one possible target being casein vesicle biogenesis in the trans-Golgi network.
Rennison
M E
ME
Department of Physiology, University of Liverpool, UK.
Handel
S E
SE
Wilde
C J
CJ
Burgoyne
R D
RD
D000818:Animals; D002364:Caseins / Q000378:metabolism; D002478:Cells, Cultured; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique; D007700:Kinetics; D007774:Lactation / Q000502:physiology; D008321:Mammary Glands, Animal / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D008854:Microscopy, Electron; D008870:Microtubules / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D015739:Nocodazole / Q000494:pharmacology*; D011506:Proteins / Q000378:metabolism*
D002364:Caseins; D011506:Proteins; D015739:Nocodazole