Figure 2.

Loss of Distal Ciliary ODA Type 2 Heavy Chains (DNAH9 and DNAH5) from the Respiratory Cilia of Individuals with DNAH9 Mutations

(A) Diagram of cilia beating in a control subject compared to a DNAH9 mutation case subject (2.II.2), showing the subtle defect in the bend of the distal portion of the cilia that is captured in HSVM (effective stroke shown in blue, recovery stroke in green). DNAH9-deficient cilia have reduced beating frequency accompanied by reduced amplitude of the distal cilia area.

(B and C) Respiratory epithelial cells were double stained with an antibody marker of the ciliary axoneme (acetylated alpha tubulin, green) and the ODA β-HC and γ-HC antibody markers (DNAH9 and DNAH5, red). Co-localization of acetylated tubulin and presence of dynein heavy chain appears yellow in merged images in the right column. Merged images of the red and green are overlaid with a light microscopy image to aid visualization of the cell and an image of the nucleus labeled with DAPI in blue.

(B) DNAH9 localizes distally in the cilia of control subject but is absent from cilia of affected individuals.

(C) In contrast to the control cilia where DNAH5 localizes along the whole length, it is highly reduced in the distal part of the cilia of affected DNAH9-mutation-carrying individuals but is still present proximally. All scale bars, 5 μM.

(D) A graph showing DNAH5 signal quantified by plotting the pixel intensity in images across profile plots of immunofluorescent images of antibody-labeled cilia, in individual 2.II.3. DNAH5 is present mainly in the proximal part of the cilia and reduced in the distal part (results from control are shown in Figure S4).