Molecular mechanisms of bleeding disorder-associated GFI1BQ287* mutation and its affected pathways in megakaryocytes and platelets.

Dominant-negative mutations in transcription factor Growth Factor Independence-1B (GFI1B) like GFI1B-Q287*, cause a bleeding disorder characterized by a plethora of megakaryocyte and platelet abnormalities. The deregulated molecular mechanisms and pathways are unknown. Here we show that both normal and Q287* mutant GFI1B interacted most strongly with the LSD1-RCOR-HDAC co-repressor complex in megakaryoblasts. Sequestration of this complex by GFI1B-Q287* and chemical separation of GFI1B from LSD1, induced abnormalities in normal megakaryocytes comparable to those seen in patients. Megakaryocytes derived from GFI1B-Q287* induced pluripotent stem cells also phenocopied abnormalities seen in patients. Proteome studies on normal and mutant induced pluripotent stem cell derived megakaryocytes identified a multitude of deregulated pathways downstream of GFI1B-Q287* including cell division and interferon signaling. Proteome studies on platelets from GFI1B-Q287* patients showed reduced expression of proteins implicated in platelet function, and elevated expression of proteins normally downregulated during megakaryocyte differentiation. Thus, GFI1B and LSD1 regulate a broad developmental program during megakaryopoiesis, and GFI1B-Q287* deregulates this program through LSD1-RCOR-HDAC sequestering.