CircFUT8 Promotes Megakaryocyte Proplatelet Formation By Regulating Actin Cytoskeleton Reorganization

Megakaryocytes (MKs) are large cells derived from hematopoietic stem cells (HSCs) through a process of differentiation and maturation. Mature MKs extend long processes called proplatelets, which are released into bone marrow sinusoids under the shear forces of blood flow. These proplatelets also form platelets in the lungs. Platelets are small cell fragments that play crucial roles in hemostasis, thrombosis, and immunity. The process of platelet generation from megakaryocytes is intricately orchestrated by non-coding RNAs, such as miRNAs. Another non-coding RNA, circRNA, is highly abundant in platelets, but its role in platelet production remains unclear. We performed RNA-seq and bioinformatics analysis to identify circRNAs during the differentiation of megakaryocytes from umbilical cord blood hematopoietic stem cells. We found that circFUT8, a novel circRNA, increases with megakaryocyte differentiation and is enriched in mature megakaryocytes. Knockdown and overexpression studies showed that circFUT8 promotes human megakaryocyte proplatelet formation (PPF) in vitro. We also found that circFUT8 is highly conserved between humans and mice. In vitro, knockdown of circFUT8 decreases mouse megakaryocyte PPF. In vivo, it reduces murine platelet counts, prolongs tail bleeding time, and reduces the number of MKs in contact with sinusoids. Furthermore, knockdown of circFUT8 reduces F-actin polymerization and impairs spreading on collagen. Additionally, we revealed that circFUT8 interacts with insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) and stabilizes tensin 1 (TNS1) mRNA. We observed that knockdown of TNS1 disrupts the actin cytoskeleton and decreases PPF. Our study highlights the crucial functions of circRNAs in platelet production and the rearrangement of the actin cytoskeleton.