Functional Genomics Approaches to Platelet Signaling

Abstract SCI-37

The count, volume and function of platelets varies in the population and there is ample evidence that all 3 quantitative traits (QT) are highly heritable. The control of platelet function is complex with a large number of forward and reverse regulatory pathways that promote and inhibit thrombus formation, respectively. We used 4 parallel genomics approaches with the aim to identify novel regulators of platelet function. First we established a genome-wide expression (GWE) HaemAtlas of all 8 haematological elements, including erythroblasts and megakaryocytes (MKs)(1). Mining of this data identified 279 transcripts that were relatively over-expressed in MKs if compared with the results obtained with the 7 remaining blood cell types. Of these, 75 transcripts encode membrane proteins, many with known and several with a hitherto unknown function (2). Second we determined the platelet response to ADP and a collagen mimetic in 500 healthy subjects of the Platelet Function Cohort (PFC)(3). WGE studies of platelet RNA samples from 37 PFC subjects, selected to be representative of the observed variation, identified 63 transcripts that were correlated with function. Third the 500 PFC samples were typed for 1536 SNPs tagging 108 candidate genes for sequence variation and this identified 19 associations (p-value ≤ 0.005) (3). All 19 QT loci (QTLs), but the GP6 gene, were novel. Based on the above 9 genes were selected for functional studies with platelets from genotyped healthy subjects and by morpholino-based gene knockdown in a model of laser-induced thrombus formation in Danio rerio (2,3). This identified BAMBI, COMMD7, LRRC32 and LRRFIP1 playing a role in the promotion and DCBLD2, ESAM, G6B and GTF2A2 in the inhibition of thrombus formation. Finally 2 of the 15 QTLs for platelet volume and count identified in about 15,000 healthy subjects with genome-wide typing information showed an effect on platelet function demonstrating that sequence variants that modify platelet volume may also exert an effect on function (4). (1) Macaulay, I. C. et al., Comparative gene expression profiling of in vitro differentiated megakaryocytes and erythroblasts identifies novel activatory and inhibitory platelet membrane proteins. Blood 109 (8), 3260 (2007); Watkins, N. A. et al., A HaemAtlas: characterizing gene expression in differentiated human blood cells. Blood 113 (19), e1 (2009); (2) O'Connor, M. N. et al., Functional genomics in zebrafish permits rapid characterization of novel platelet membrane proteins. Blood 113 (19), 4754 (2009).; (3) Jones, C. I. et al., A functional genomics approach reveals novel quantitative trait loci associated with platelet signaling pathways. Blood 114 (7), 1405 (2009); (4) Soranzo, N. et al., A genome-wide meta-analysis identifies 22 loci associated with eight hematological parameters in the HaemGen consortium. Nat Genet 41 (11), 1182 (2009).