Cell-autonomous FLT3L shedding via ADAM10 mediates conventional dendritic cell development in mouse spleen

Abstract

Conventional dendritic cells (cDCs) derive from bone marrow (BM) precursors that undergo cascades of developmental programs to terminally differentiate in peripheral tissues. Pre-cDC1s and pre-cDC2s commit in the BM to each differentiate into CD8α+/CD103+ cDC1s and CD11b+ cDC2s, respectively. Although both cDCs rely on the cytokine FLT3L during development, mechanisms that ensure cDC accessibility to FLT3L have yet to be elucidated. Here, we generated mice that lacked a disintegrin and metalloproteinase (ADAM) 10 in DCs (Itgax-cre × Adam10-fl/fl; ADAM10∆DC) and found that ADAM10 deletion markedly impacted splenic cDC2 development. Pre-cDC2s accumulated in the spleen with transcriptomic alterations that reflected their inability to differentiate and exhibited abrupt failure to survive as terminally differentiated cDC2s. Induced ADAM10 ablation also led to the reduction of terminally differentiated cDC2s, and restoration of Notch signaling, a major pathway downstream of ADAM10, only modestly rescued them. ADAM10∆DC BM failed to generate cDC2s in BM chimeric mice with or without cotransferred ADAM10-sufficient BM, indicating that cDC2 development required cell-autonomous ADAM10. We determined cDC2s to be sources of soluble FLT3L, as supported by decreased serum FLT3L concentration and the retention of membrane-bound FLT3L on cDC2 surfaces in ADAM10∆DC mice, and by demonstrating the release of soluble FLT3L by cDC2 in ex vivo culture supernatants. Through in vitro studies utilizing murine embryonic fibroblasts, we determined FLT3L to be a substrate for ADAM10. These data collectively reveal cDC2s as FLT3L sources and highlight a cell-autonomous mechanism that may enhance FLT3L accessibility for cDC2 development and survival.

TAO-kinase 3 governs the terminal differentiation of NOTCH2-dependent splenic conventional dendritic cells

“…Yet, the drastic reduction in NOTCH2dependent cDC1s and cDC2s in conditional ADAM10 knockout mice strongly suggests that ADAM10 is the essential NOTCH2 sheddase in DCs. In support of our findings, a role of dendritic cell ADAM10 in the differentiation of cDC2s and the generation of T H 2 responses was previously reported by us and others (53,54). Although off-target effects of the CD11c-driven Cre have been described (55,56), our mixed bone marrow experiments clearly confirmed the effects of ADAM10 to be cell-intrinsic.…”

Transcriptional network analysis of transcriptomic diversity in resident tissue macrophages and dendritic cells in the mouse mononuclear phagocyte system

“…[96, 104,105]. Consistent with the literature, Itgax was expressed in multiple macrophage populations (Figure 2A) at levels at least as high as in purified DC, and correlated only with Cd22, Cd274 (encoding PD-L1), Csf2rb, Csf2rb2, Slc15a3, Tmem132a and the transcription factor gene Prdm1 (Cluster145).…”

“…The transcript encoding the ligand (Flt3l) was expressed constitutively to varying degrees in all of the MPS populations studied. Fujita et al [104] showed that FLT3L is cleaved from the cell surface of expressing cells by ADAM10. Conditional deletion of Adam10 using Itgax-cre led to reduced differentiation of cDC2.…”

Network analysis of transcriptomic diversity amongst resident tissue macrophages and dendritic cells in the mouse mononuclear phagocyte system

“…Despite evidence that it is expressed by many resident tissue macrophages (reviewed in [ 25 ]), CD11C (encoded by Itgax ) is still widely used as a surface marker in mouse DC purification. Ongoing studies of the impacts of conditional mutations using Itgax -cre continue to be interpreted solely in terms of DC specificity (for example, [ 121 , 129 , 130 ]). Consistent with the literature [ 25 ], Itgax was expressed in multiple macrophage populations other than DCs ( Fig 2A ) at levels at least as high as in DCs purified using CD11C as a marker and correlated only with Cd22 , Cd274 (encoding programmed cell death 1 ligand 1), Csf2rb , Csf2rb2 , solute carrier ( Slc ) 15a3 , Tmem132a , and the transcription factor gene Prdm1 (Cluster145).…”

“…The transcript encoding the FLT3 ligand ( Flt3l ) was expressed constitutively to varying degrees in all of the MPS populations studied. Fujita and colleagues [ 129 ] showed that FLT3L is cleaved from the cell surface of expressing cells by ADAM10. Conditional deletion of Adam10 using Itgax -cre led to reduced differentiation of cDC2s.…”