(PLEASE KEEP IN WORD, DO NOT PDF) Title: The role of Septin6 Group in Murine and Human Hematopoiesis 1 R01 DK137172-01 Septins are highly conserved GTPase proteins which regulate a variety of cellular functions including cytokinesis, polarity, cell cycle, vesicle trafficking, exocytosis, and creation of diffusion barriers. We have recently identified a non-syndromic newborn with severe neutropenia (Renella, AJH 2022) with a novel X-linked germline mutation in the C-terminus of SEPTIN6 gene (SEPTIN6 c.1282T>C p.428Glnext*9) associated with dysmyelopoiesis. Editing the germline mutation into normal male CD34+ hematopoietic stem and progenitor cells (HSC/P) phenocopied key pathologic features of the clinical syndrome, including large, multinucleated nuclei and reduced myeloid progenitor growth in vitro (preliminary data). The C-terminus has been proposed to play a key role in filament stabilization, bundling and bending as well as interactions with other septins and in silico modeling suggest the addition of 9 amino acids (aa) associated with the SEPTIN6 c.1282T>C p.428Glnext*9 mutation would alter function by interfering with SEPTIN6 protein interactions. Based on our initial publication, a second patient with severe neutropenia has since been identified in Seattle with a distinct mutation in the same codon (SEPTIN6 c.1282T>A) (Mohamad, Allensbach, Williams, accepted for presentation, ASH 2022). While the aa substitution is different, modeling predicts that SEPTIN6 variants adopt the same structure. Both identified patients had a high degree of myeloid tetraploidy in the marrow and were refractory to G-CSF.

Project Narrative Our studies will define the role(s) of septin6 group proteins in murine and human hematopoiesis with a focus on the unique versus overlapping roles of Septin6, Septin8, and Septin11, specifically in hematopoietic cell migration and homing, differentiation, proliferation, and engraftment and correlate these phenotypes with cytokinesis, cell cycle, polarity, apoptosis and the aging phenotype. We will investigate the nature of the abnormal blood and marrow findings in humans with mutations of SEPTIN6 that we have recently described.