41003:

Dynamic reorganization of endoplasmic reticulum along actin filaments and microtubules.

Authors:	Marc, Jan(A)Granger, Cheryl, L.(B)Cyr, Richard , J.(B)
Affiliations:	(A): Biological Sciences, University of Sydney (B): Department of Biology, Pennsylvania State University
Presenter:	Marc, Jan , jmarc@bio.usyd.edu.au

Endoplasmic reticulum (ER), the first compartment in the protein synthesis machinery, is a complex network of lamellar and tubular cisternae of unique morphologies and functions. In plant cells, the membranous network generally extends throughout the cytoplasm and undergoes dynamic rearrangements. Previous kinetic analysis using the fluorescent probe DiOC6 showed that ER motility is driven by acto-myosin motor associated with cytoplasmic streaming. Details of the mechanism, however, are unknown. We have introduced an ER-targetted GFP construct (courtesy of Jim Haseloff, MRC Laboratory of Molecular Biology, Cambridge) into the lower epidermis of bean leaves, and studied the reorganization of the ER in large epidermal cells along the midvein using epifluorescence microscopy and a CCD camera. In addition to massive streaming at a velocity of 2-10 mm/s along the large endoplasmic cables, the ER also travelled in finer streams in the cell cortex amongst a relatively immobile network of ER sheets and windows with polygonal partitions of tubular ER. Interestingly, the cortical streaming was usually oriented at transverse or oblique angles relative to the cell length. Application of Cytochalasin D stopped the endoplasmic mainstream within a few minutes and converted it into amorphous stationary clumps, while the tubular ER in the cortex formed oblong windows oriented at distinct transverse or oblique angles, reminiscent of the typical transverse or oblique orientation of cortical microtubules. Application of the microtubule drugs ami-prophos methyl (APM) or taxol had no effect on the endoplasmic mainstream. However, APM distorted the cortical directional streaming and also prevented the formation of the transverse or oblique windows of tubular ER when applied in combination with Cytochalasin D, suggesting that microtubules are involved in directing the streaming in the cortical cytoplasm. To confirm this hypothesis, we co-transformed epidermal cells with the ER-GFP together with a GFP-microtubule binding reporter gene (Marc et al 1998, Plant Cell 10: 1927-1939). This showed that the transverse or oblique cortical streaming does indeed coincide with the location of cortical microtubules, and hence that the acto-myosin and microtubule cytoskeletons are interconnected and both support rearrangements of the ER. Supported by ARC grant A19700148 to JM.