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. 2020 Jan 23;106(2):153–169. doi: 10.1016/j.ajhg.2019.12.010

Figure 3.

Figure 3

TTC12 Loss-of-Function Mutations Cause Defective Single-Headed IDA Assembly in Respiratory Cilia

(A) Protein lysates extracted from AECs of a control individual and individuals 18GM00157, DCP1606, and DCP153 were resolved by SDS-PAGE and analyzed by immunoblotting with an anti-TTC12 antibody. The expression of TTC12 is virtually absent or dramatically reduced in PCD-affected individuals. DNAJB13, a component of radial spokes, was used as a positive control.

(B) IF microscopy analysis of IDA markers in respiratory epithelial cells from a control and from individuals 18GM00157, DCP1606, and DCP153 homozygous for TTC12 mutations. Cells were double-labeled with an anti-acetylated α-tubulin antibody (red), a marker of ciliary axonemes, and anti-DNAH6, anti-DNAH12, or anti-DNALI1 antibodies as markers of IDAs (green). Nuclear staining was performed with DAPI (blue). In affected individuals, DNAH12 is completely absent in the cilia but seems to accumulate at the subapical part of the cytoplasm. By contrast, DNALI1 and DNAH6 appear to be strongly reduced but are partially present at the proximal part of cilia. (DNAH2 and DNAH10, two markers of the double-headed IDA-I1, were shown to be normally expressed in the individuals; see Figure S7). Scale bars represent 5 μm. IDA: inner dynein arm.