Original Article

In Vitro Formation of the Endoplasmic Reticulum Occurs Independently of Microtubules by a Controlled Fusion Reaction

Correspondence to: Tom A. Rapoport, Howard Hughes Medical Institute and Department of Cell Biology, Harvard Medical School, 240 Longwood Ave., Boston, MA 02115-6091. Tel:(617) 432-0637 Fax:(617) 432-1190 E-mail:tom_rapoport{at}hms.harvard.edu.

We have established an in vitro system for the formation of the endoplasmic reticulum (ER). Starting from small membrane vesicles prepared from Xenopus laevis eggs, an elaborate network of membrane tubules is formed in the presence of cytosol. In the absence of cytosol, the vesicles only fuse to form large spheres. Network formation requires a ubiquitous cytosolic protein and nucleoside triphosphates, is sensitive to N-ethylmaleimide and high cytosolic Ca²⁺ concentrations, and proceeds via an intermediate stage in which vesicles appear to be clustered. Microtubules are not required for membrane tubule and network formation. Formation of the ER network shares significant similarities with formation of the nuclear envelope. Our results suggest that the ER network forms in a process in which cytosolic factors modify and regulate a basic reaction of membrane vesicle fusion.

Key Words: membrane tubules, endoplasmic reticulum, membrane fusion, microtubules, Xenopus

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