Insights into megakaryocyte biology from population platelet traits

Abstract

Platelets are anucleate blood cells released from bone marrow megakaryocytes (MKs) that are essential mediators of haemostasis. Genome-wide association studies (GWAS) of platelet count and size measured by clinical haematology analysers have already identified informative platelet genes. GWAS may also be applied to traits such as platelet side scatter (PLT SSC), a surrogate for cytoplasmic complexity. Pre-publication access to data from collaborators from the first ever GWAS for PLT SSC involving 39,656 INTERVAL study blood donors provided a unique opportunity to identify new genetic associations. The overarching aim of this research was to use these data to identify and functionally evaluate candidate genes associated with PLT SSC to gain biological insights into platelet and MK biology.

PLT SSC was found to be a heritable trait that was distinct but correlated with platelet size. Statistical and experimental evidence indicated that PLT SSC was a marker of platelet activation, that in a phenome-wide analysis was associated with inflammatory disorders, breast cancer and COVID-19 severity.

Integration of conditionally significant variants for PLT SSC adjusted for other platelet parameters (ADJ PLT SSC) with population genomic datasets identified candidate genes, including ZFPM2 which encodes a GATA transcription cofactor. ZFPM2 was associated with multiple platelet traits and colocalised with plasma pQTLs for platelet α-granule proteins and thrombotic disease. Individuals with predicted pathogenic variants in ZFPM2 tended to have lower PLT and higher MPV.

Ribonucleoprotein-based CRISPR-Cas9 gene editing of ZFPM2 in imMKCL cells reduced proliferation of MK progenitors, likely through cell cycle arrest, and impaired nuclear and cytoplasmic maturation. Potential mechanisms were explored using RNA sequencing. Platelets generated from ZFPM2 targeted imMKCL cells showed a trend towards larger size and SSC, consistent with the initial GWAS findings. This research highlights the power of GWAS to identify hitherto uncharacterised regulatory pathways in blood cells.

Date of Award	3 Oct 2023
Original language	English
Awarding Institution	University of Bristol
Supervisor	Alastair W Poole (Supervisor), Andrew D Mumford (Supervisor) & Jonathan Mill (Supervisor)