VAMP8/endobrevin is overexpressed in hyperreactive human platelets: suggested role for platelet microRNA.
BACKGROUND
Variation in platelet reactivity contributes to disorders of hemostasis and thrombosis, but the molecular mechanisms are not well understood.


OBJECTIVE
To discover associations between interindividual platelet variability and the responsible platelet genes, and to begin to define the molecular mechanisms altering platelet gene expression.


METHODS
Two hundred and eighty-eight healthy subjects were phenotyped for platelet responsiveness. Platelet RNA from subjects demonstrating hyperreactivity (n=18) and hyporeactivity (n=11) was used to screen the human transcriptome.


RESULTS
Distinctly different mRNA profiles were observed between subjects with differing platelet reactivity. Increased levels of mRNA for VAMP8/endobrevin, a critical v-SNARE involved in platelet granule secretion, were associated with platelet hyperreactivity (Q=0.0275). Validation studies of microarray results showed 4.8-fold higher mean VAMP8 mRNA levels in hyperreactive than hyporeactive platelets (P=0.0023). VAMP8 protein levels varied 13-fold among platelets from these normal subjects, and were 2.5-fold higher in hyperreactive platelets (P=0.05). Among our cohort of 288 subjects, a VAMP8 single-nucleotide polymorphism (rs1010) was associated with platelet reactivity in an age-dependent manner (P<0.003). MicroRNA-96 was predicted to bind to the 3'-untranslated regionof VAMP8 mRNA and was detected in platelets. Overexpression of microRNA-96 in VAMP8-expressing cell lines caused a dose-dependent decrease in VAMP8 protein and mRNA, suggesting a role in VAMP8 mRNA degradation.


CONCLUSIONS
These findings support a role for VAMP8/endobrevin in the heterogeneity of platelet reactivity, and suggest a role for microRNA-96 in the regulation of VAMP8 expression.
Kondkar
A A
AA
Thomas Jefferson University, The Cardeza Foundation for Hematologic Research and the Department of Medicine, Jefferson Medical College, Philadelphia, PA 19107, USA.
Bray
M S
MS
Leal
S M
SM
Nagalla
S
S
Liu
D J
DJ
Jin
Y
Y
Dong
J F
JF
Ren
Q
Q
Whiteheart
S W
SW
Shaw
C
C
Bray
P F
PF
D020413:3' Untranslated Regions; D000328:Adult; D000367:Age Factors; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism*; D004837:Epinephrine; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods; D005838:Genotype; D045325:HCT116 Cells; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000097:blood*; D020411:Oligonucleotide Array Sequence Analysis; D010641:Phenotype; D010974:Platelet Aggregation / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D050683:R-SNARE Proteins / Q000097:blood / Q000235:genetics*; D012333:RNA, Messenger / Q000097:blood; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D015854:Up-Regulation; D055815:Young Adult
D020413:3' Untranslated Regions; C541492:MIRN96 microRNA, human; D035683:MicroRNAs; D050683:R-SNARE Proteins; D012333:RNA, Messenger; C112938:VAMP8 protein, human; D004837:Epinephrine