MEMBRANE/MICROTUBULE TIP ATTACHMENT COMPLEXES (TACS) ALLOW THE ASSEMBLY DYNAMICS OF PLUS ENDS TO PUSH AND PULL MEMBRANES INTO TUBULOVESICULAR NETWORKS IN INTERPHASE XENOPUS EGG EXTRACTS

We discovered by using high resolution video microscopy, that membranes become attached selectively to the growing plus ends of microtubules by membrane/microtubule tip attachment complexes (TACs) in interphase-arrested, undiluted, Xenopus egg extracts, Persistent plus end growth of stationary microtubules pushed the membranes into thin tubules and dragged them through the cytoplasm at the similar to 20 mu m/min velocity typical of free plus ends. Membrane tubules also remained attached to plus ends when they switched to the shortening phase of dynamic instability at velocities typical of free ends, 50-60 mu m/min. Over time, the membrane tubules contacted and fused with one another along their lengths, forming a polygonal network much like the distribution of ER in cells. Several components of the membrane networks formed by TACs were identified as ER by immunofluorescent staining using antibodies to ER-resident proteins. TAC motility was not inhibited by known inhibitors of microtubule motor activity, including 5 mM AMP-PNP, 250 mu M orthovanadate, and ATP depletion. These results show that membrane/microtubule TACs enable polymerizing ends to push and depolymerizing ends to pull membranes into thin tubular extensions and networks at fast velocities.