A tip-high gradient of a putative plasma membrane SNARE approximates the exocytotic gradient in hyphal apices of the fungus Neurospora crassa.
Antibodies to the Saccharomyces cereviseae plasma membrane t-SNARE Sso2p identify a putative 39-kDa homologue in Neurospora crassa. The 39-kDa protein is enriched in plasma membrane (PM) and occurred with other membranes. It is extractable by detergent, but not chaotropic or alkali agents, suggesting membrane insertion. Immunoprecipitation with anti-Sso2p coprecipitated a approximately 100-kDa, Mg(+)-ATP-sensitive band with the 39-kDa protein, suggesting a ternary SNARE complex. Affinity-purified anti-Sso2p gave hyphal staining patterns most consistent with protein localization on both the PM and intracellular exocytotic apical wall vesicles. The PM staining in hyphal apices formed a tip-high gradient, not as steep as that predicted by the "hyphoid equation," but closer to published gradients of cell wall matrix deposition. We conclude that the t-SNAREs are transported to the PM on the apical vesicles, but their tip-high gradient alone is insufficient to explain the vesicle fusion gradient in growing tips.
Gupta
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GD
Department of Biology, York University, 4700 Keele Street, Toronto, Ontario, M3J 1P3, Canada.
Heath
I B
IB
D005089:Exocytosis; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D015151:Immunoblotting; D007150:Immunohistochemistry; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009492:Neurospora crassa / Q000254:growth & development* / Q000378:metabolism*; D011233:Precipitin Tests; D050765:Qa-SNARE Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins
D005656:Fungal Proteins; D008565:Membrane Proteins; D050765:Qa-SNARE Proteins; C084007:SSO2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins