Reorganization of the Secretory Pathway:
In addition to the assembly of a meiotic outer plaque, formation of the prospore membrane requires a reorganization of the secretory pathway so that exocytic vesicles are delivered to the intracellular prospore membrane rather than the plasma membrane. The mechanism by which the vesicles are retargeted is unknown. As well as the redirection of exocytic vesicles other changes in the secretory pathway are necessary, including a requirement for the phospholipase D enzyme encoded by SPO14 and a requirement for a sporulation-specific SNARE protein, Spo20p.
Figure: Immunfluorescence image showing the localization of the plasma membrane t-SNARE Sso1p to the prospore membrane in sporulating cells. Left panel, anti-Sso1p staining. Right panel, DAPI staining of corresponding cells.
SNAREs are heterooligomeric complexes that mediate the fusion between vesicles and membranes within the cell. Fusion of exocytic vesicles with the yeast plasma membrane requires a complex that consists of the Snc protein, the Sso protein, and the Sec9 protein. For fusion of exocytic vesicles with the prospore membrane, Sec9p is replaced by a related protein, Spo20p. Although Sec9 and Spo20 are 40% identical and form similar complexes with Sso1p and Snc1p, their ability to mediate vesicle fusion is largely restricted to the plasma membrane and the prospore membrane, respectively. The basis for this restriction is actively under investigation.
Figure: An analysis of Spo20p/Sec9p chimeric molecules reveals two regions of the Spo20p protein critical for function.
The switch between Sec9p and Spo20p mediated membrane fusion may represent some underlying difference in the protein or lipid compositions of the plasma and prospore membranes. In fact, Spo20p seems to have a preference in vivo for membranes with higher levels of phosphatidic acid (PA). Increasing PA levels in the plasma membrane will allow Spo20p to rescue a sec9 mutant. Also, a region in the amino-terminus of Spo20p binds to PA in vitro, is essential for function, and is sufficient to localize the protein to the prospore membrane in vivo.
Figure: Fusion of GFP to aa51-91 of Spo20p localizes GFP to the prospore membrane in sporulating cells. b,c,d; GFP-Spo20p51-91. f,g,h DAPI staining of chromatin in the same cells
| For More Details on our work in this area see references #1, #4, #10, #11 |
|
Additional reading: Rudge, S.A., Morris, A.J.,& J. Engebrecht (1998) Relocalization of phospholipase D activity mediates membrane formation during meiosis. J Cell Biol. 140:81-90. [PubMed] Jantti, J., Aalto, M. K., Oyen, M., Sundqvist, L., Keranen, S., &
H. Ronne (2002) Characterization of temperature-sensitive mutations in
the yeast syntaxin 1 homologues Sso1p and Sso2p, and |