Investigation of factors regulating von Willebrand factor expression

Summary The overall aim of this project is to characterize the non-coding genomic sequences involved in regulating the transcriptional activity at the von Willebrand factor locus (VWF). Deficiencies and supranormal levels of VWF are associated with bleeding and thrombotic disorders, respectively. However, the mechanistic basis for determining VWF plasma levels, both in healthy and quantitative pathologic state remains poorly understood. Three specific aims are described in this project. In Aim #1, we will investigate details of the transcriptional regulation of VWF and FVIII in various subtypes of endothelium. This will involve a combination of experimental and computational studies including single cell analyses and spatial transcriptomic investigation. The results of these studies will provide new information concerning the cell types, transcriptomic pathways, and anatomic niches involved in the expression of these proteins. In Aim #2, we will investigate the non-coding genomic sequences engaged in the basal transcriptional regulation of VWF. These studies will utilize a range of orthogonal in-silico and cell-based strategies to evaluate the regulatory potential of non-coding candidate regions both within and adjacent to the VWF locus. The information derived from these studies will subsequently be integrated into the analysis of the population cohort studies of subjects with low and high levels of VWF enrolled in Projects 1 & 2 of this PPG. Lastly, in Aim #3, we will conduct studies to address the phenomenon of the age-dependent increase in plasma VWF. We have evidence from prior mouse studies that increased VWF mRNA production contributes significantly to this phenomenon. In Aim #3 we will evaluate several non-coding sequences for which we have preliminary results indicating biological plausibility for their causal engagement in age-dependent VWF expression. These studies will utilize in-silico and various cell- based approaches including the assessment of transcriptional activity in conditions of shear-flow and inflammation simulating the aging vasculature. Lastly, further studies will be conducted in aging mice to provide complementary in vivo support for our in vitro findings.