Deachon, Karen A.
SNARE specificity in membrane protein traffic.
MSc(R) thesis, University of Glasgow.
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Recent findings indicate that there is a role for SNAREs in ion channel regulation. With the discovery of the vesicle trafficking SNARE protein NtSYP121 came the development of the predicted cytosolic domain fragment of the protein. This Sp2 fragment was found in studies to block the responses of guard cell K+ and Cl- channels to ABA. These observations raised questions about the role of SNAREs in ion channel control. In this study, the effects of several plasma membrane SNARE Sp2s and one PVC-localised SNARE Sp2 on the K+ channel KAT1 were investigated. The experiments made use of GFP- and HA- tagged KAT1 to observe the traffic and localisation of KAT1 in tobacco cells. Acetylsalicylic acid was also used to combat the effects of the wound response activated during slide preparation. Coexpression of KAT1 with SYP121-Sp2, SYP122-Sp2 and SYP71-Sp2 resulted in a disruption of trafficking and in microdomains becoming diffuse and mobile at the plasma membrane. No disruption in trafficking or localisation of KAT1 was seen when coexpressed with the PVC-localised SNARE fragment SYP21-Sp2. None of the Sp2s used had any effect on the H+-ATPase PMA2. These results offer evidence of a role for plasma membrane SNAREs in trafficking and anchoring of KAT1 to the plasma membrane.
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