In situ synthesis of beta-glucan microfibrils on tobacco plasma membrane sheets

A major concern in plant morphogenesis is whether cortical microtubules are responsible for the arrangement and action of beta-glucan synthases in the plasma membrane. We prepared isolated plasma membrane sheets with cortical microtubules attached and tested whether beta-glucan synthases penetrated through the membrane to form microfibrils and whether these synthases moved in the fluid membrane along the cortical microtubules. This technique enabled us to examine synthesis of beta-glucan as a fiber with a two-dimensional structure. The synthesis of beta-glucan microfibrils was directed in arrays by cortical microtubules at many loci on the membrane sheets. The microfibrils were mainly arranged along the microtubules, but the distribution of microfibrils was not always parallel to that of the microtubules. The rate of beta-glucan elongation as determined directly on the exoplasmic surface was 620 nm per min. When the assembly of microtubules was disrupted by treatment with propyzamide, the beta-glucans were not deposited in arrays but in masses. This finding shows that the arrayed cortical microtubules are not required for beta-glucan synthesis but are required for the formation of arranged microfibrils on the membrane sheet.