Nbeal2 interacts with Dock7, Sec16a, and Vac14.

Mutations in NBEAL2, the gene encoding the scaffolding protein Nbeal2, are causal of gray platelet syndrome (GPS), a rare recessive bleeding disorder characterized by platelets lacking α-granules and progressive marrow fibrosis. We present here the interactome of Nbeal2 with additional validation by reverse immunoprecipitation of Dock7, Sec16a, and Vac14 as interactors of Nbeal2. We show that GPS-causing mutations in its BEACH domain have profound and possible effects on the interaction with Dock7 and Vac14, respectively. Proximity ligation assays show that these 2 proteins are physically proximal to Nbeal2 in human megakaryocytes. In addition, we demonstrate that Nbeal2 is primarily localized in the cytoplasm and Dock7 on the membrane of or in α-granules. Interestingly, platelets from GPS cases and Nbeal2-/- mice are almost devoid of Dock7, resulting in a profound dysregulation of its signaling pathway, leading to defective actin polymerization, platelet activation, and shape change. This study shows for the first time proteins interacting with Nbeal2 and points to the dysregulation of the canonical signaling pathway of Dock7 as a possible cause of the aberrant formation of platelets in GPS cases and Nbeal2-deficient mice.

Perivascular niche cells sense thrombocytopenia and activate hematopoietic stem cells in an IL-1 dependent manner.

Hematopoietic stem cells (HSCs) residing in specialized niches in the bone marrow are responsible for the balanced output of multiple short-lived blood cell lineages in steady-state and in response to different challenges. However, feedback mechanisms by which HSCs, through their niches, sense acute losses of specific blood cell lineages remain to be established. While all HSCs replenish platelets, previous studies have shown that a large fraction of HSCs are molecularly primed for the megakaryocyte-platelet lineage and are rapidly recruited into proliferation upon platelet depletion. Platelets normally turnover in an activation-dependent manner, herein mimicked by antibodies inducing platelet activation and depletion. Antibody-mediated platelet activation upregulates expression of Interleukin-1 (IL-1) in platelets, and in bone marrow extracellular fluid in vivo. Genetic experiments demonstrate that rather than IL-1 directly activating HSCs, activation of bone marrow Lepr+ perivascular niche cells expressing IL-1 receptor is critical for the optimal activation of quiescent HSCs upon platelet activation and depletion. These findings identify a feedback mechanism by which activation-induced depletion of a mature blood cell lineage leads to a niche-dependent activation of HSCs to reinstate its homeostasis.