Title:
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Identification and functional
characterisation of novel SNARE
proteins in platelets
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Platelet secretion not only drives thrombosis and haemostasis, but also mediates
a variety of other physiological and pathological processes. The ubiquitous
SNARE machinery and a number of accessory proteins have been implicated in
regulating secretion in platelets. Although several platelet SNAREs have been
identified, further members of the SNARE family may help fine-tune platelet secretion.
In this study I identified expression of t-SNAREs VTIlA, VTIlB (Qb
SNAREs) and STX8 (Qc SNARE) in human and mouse platelets. Those 'novel'
SNAREs were able to interact with each other and previously reported SNAREs
VAMP8 (R-SNARE) and STXll (Qa SNARE), thus suggesting existence of a
secondary SNARE complex in addition to the widely accepted SNAP23-VAMP8-
STXll complex. In following mouse studies, whereas neither gene was found to
be essential for a -granule or lysosome secretion, Stx8-/- platelets showed a
significant defect in dense granule secretion and aggregation, that was most
pronounced at intermediate concentrations of agonists. Addition of exogenous
ADP could rescue the aggregation defect but failed to restore dense granule
secretion, suggesting the defect lies in the 'primary' secretory pathway. Pretreatment
with P2Y receptors antagonists reduced secretion and aggregation to
the same extent in WT and Stx8-/- platelets, suggesting that the ADP-driven
positive feedback mechanism was not defective in Stx8-/- platelets. In addition,
STX8 was found to play a role in regulating pro-coagulant activity suggesting
novel roles for SNAREs in platelets. Neither Vtila-/- nor Vtilb-/- showed any
significant defects, suggesting complementarity between those homologues in
platelets. STX8 therefore specifically contributes to dense granule secretion and
represents another member of a growing family of genes that play distinct roles
in differentially regulating granule release from platelets. Taken together, data
presented in this Thesis not only provides first evidence of an additional SNARE
complex present in platelets but also suggests novel roles for SNAREs in
regulation of thrombosis and haemostasis.
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