To gain insight into the spatiotemporal expression of the different ODA HC-proteins in human respiratory cilia, we then analyzed the localization of both β-ODA HCs DNAH9 and DNAH11 along the ciliary axonemes during ciliogenesis in control cells. Primary respiratory cell cultures and in vitro ciliogenesis experiments were performed as described in the Supplemental Methods. In early ciliogenesis stages (day 4–8) only DNAH11, but not DNAH9, is assembled into the ciliary axoneme. Between ciliogenesis day 10 and 12, we then observed localization of DNAH9 in the very distal ciliary axoneme, which extended during ciliogenesis (day 14) (Figure S10). Interestingly, Oltean et al. showed also an asynchronous expression/localization of both β-ODA HCs during ciliogenesis of respiratory cilia consistent with our findings.47 In contrast, in DNAH11 mutant cilia, DNAH9 is already present in short ciliary axonemes suggesting assembly at early stages of ciliogenesis and localization was panaxonemal in all analyzed ciliary axonemes independent of the cilia length (Figures 5, S11, and S12).6 On the other hand, proximal DNAH11 localization was not altered in DNAH9, DNAI2, or DNAH5 mutant respiratory cilia as well as DNAAF2 and DNAAF4 mutant respiratory cells (MIM: 612517 and 608706, respectively), indicating that DNAH11 evolved as a specific proximal β-ODA HC (Figures S13 and S14). Localization of the ODA docking complex proteins CCDC114 and TTC25 and the N-DRC component GAS8 was not affected in DNAH9-deficient ciliary axonemes (Figures 4B and S15). Thus, the assembly of the ODA docking complex and the N-DRC does not depend on DNAH9 function.