Heterozygous nonsense FLI1Met100* variant results in thrombocytopenia and increased erythroid gene expression

ABSTRACT: Megakaryocytes and platelets arise from hematopoietic stem and progenitor cells through a tightly regulated process involving various transcription factors, including ETS family member FLI1. Pathogenic variants in FLI1, particularly those affecting the ETS DNA-binding domain, have been implicated in inherited thrombocytopenia and are associated with abnormal giant alpha granules, as seen in Paris-Trousseau Syndrome. However, the precise pathophysiological mechanisms remain unclear. Here, we describe a patient with a de novo heterozygous nonsense mutation, FLI1Met100* (c.297del), and investigate its effects on megakaryopoiesis using the Meg01 megakaryoblast cell line as a model. We hypothesized that this variant generates a truncated protein that exerts a dominant-negative effect on megakaryocyte maturation. Western blot analysis confirmed the presence of the truncated protein following FLI1Met100* overexpression in Meg01 cells. To further examine its impact, we overexpressed both wild-type FLI1 (FLI1WT) and FLI1Met100* in Meg01 cells, followed by bulk RNA sequencing and proteomics analysis. Gene set enrichment analysis revealed that FLI1WT enhanced megakaryocytic phenotypes while suppressing erythroid phenotypes, whereas FLI1Met100* upregulated both. Flow cytometry further confirmed that erythroid markers CD235a, CD36, and KLF1 were downregulated by FLI1WT but elevated by FLI1Met100*. Additionally, FLI1WT promoted megakaryocyte maturation and adhesion, as evidenced by increased expression of the megakaryocyte marker CD61 and adhesion markers CD34 and CD44. Moreover, we demonstrated through immunoprecipitation that both FLI1WT and FLI1Met100* interact with shared cofactors. Collectively, our findings suggest that FLI1Met100* impairs megakaryocyte maturation through a dominant-negative manner, potentially contributing to thrombocytopenia by promoting erythroid features at the expense of proper megakaryocytic development.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Megakaryocyte, Cell Culture

SUBMITTER: Arie Hoogendijk  

LAB HEAD: A.J. Hoogendijk

PROVIDER: PXD063984 | Pride | 2025-12-05

REPOSITORIES: Pride