Abstract
By crossing septin7-floxed mice with Lyz2-Cre mice carrying the Cre recombinase inserted in the Lysozyme-M (Lyz2) gene locus we aimed the specific deletion of septin7 in myeloid cells, such as monocytes, macrophages and granulocytes. Septin7 flox/flox :Lyz2-Cre mice show no alterations in the myeloid compartment. Septin7-deleted macrophages (BMDMs) were isolated and analyzed. The lack of Septin7 expression was confirmed and a constitutive double-nucleation was detected in Septin7-deficient BMDMs indicating a defect in macrophage cytokinesis. However, phagocytic function of macrophages as judged by uptake of labelled E. coli particles and LPS-stimulated macrophage activation as judged by induction of TNF mRNA expression and TNF secretion were not compromised. In addition to myeloid cells, Lyz2-Cre is also active in type II pneumocytes (AT2 cells). We monitored lung adenocarcinoma formation in these mice by crossing them with the conditional knock-in Kras-LSL-G12D allele. Interestingly, we found that control mice without septin7 depletion die after 3–5 weeks, while the Septin7-deficient animals survived 11 weeks or even longer. Control mice sacrificed in the age of 4 weeks display a bronchiolo-alveolar hyperplasia with multiple adenomas, whereas the Septin7-deficient animals of the same age are normal or show only a weak multifocal brochiolo-alveolar hyperplasia. Our findings indicate an essential role of Septin7 in macrophage cytokinesis but not in macrophage function. Furthermore, septin7 seems absolutely essential for oncogenic Kras-driven lung tumorigenesis making it a potential target for anti-tumor interventions.
Highlights
Septins, a conserved family of filament forming GTPases, build heteropolymeric higher-order structures and participate in diverse cellular processes, and are being widely accepted as the fourth component of eukaryotic cytoskeleton (Mostowy and Cossart, 2012)
Even though it was a known fact that septins are required for the proliferation of several tumor-derived epithelial cell lines and Septin7-deficient fibroblasts undergo obligate multinucleation in vitro, this is the first report establishing a clear role for septins in tumorigenesis in vivo
It should be noted that the complete depletion of Septin7 in majority of the bone marrow-derived macrophages (BMDMs) was only achieved upon prolonged differentiation of bone marrow (BM) cells with MCSF
Summary
A conserved family of filament forming GTPases, build heteropolymeric higher-order structures and participate in diverse cellular processes, and are being widely accepted as the fourth component of eukaryotic cytoskeleton (Mostowy and Cossart, 2012). Septin7-deleted CD8+ T cells display cytokinetic failure upon cytokine stimulation in the absence of antigen-presenting cells (Menon et al, 2014; Mujal et al, 2016). Septin7-deficient myeloid progenitors are capable of colony formation in vitro (Menon et al, 2014) and siRNAmediated Septin depletion in myeloid K562 cell line had no impact on cell proliferation (Sellin et al, 2011). While most of the studies investigating the role of septins in bacterial pathogenesis have been performed in non-phagocytic cells, an infection model in zebrafish has shown a role for macrophages and neutrophil septins in limiting Shigella infection (Mostowy et al, 2013). We crossed the Septin7flox mice with the myeloidspecific Lyz2-Cre line (Clausen et al, 1999) to generate a tissuespecific Septin7-knockout (KO) mice and to investigate the role of septins in myeloid cells
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