Figure 3. Loss of TALPID3 (KIAA0586) causes abnormal tissue and cell polarity and abnormal intracellular organization.
(A, A′, A′′) β-catenin expression is localized anteriorly within feather buds of the wildtype chicken at day 9.5 and (B, B′, B′′) in the talpid3 chicken at day 9.5. Black circles indicate feather buds with correct polarity; dashed black circles represent no polarity; blue circles represent abnormal polarity (Schematic C). The talpid3 chicken (B′′) demonstrates feather buds which lack polarity (blue circles) which is not seen in the wildtype chicken (A′′). Asterisks represent merged feather buds. (C′) Quantification of the percentage of feather buds with correct, abnormal or no polarity in wildtype and talpid3 (D, E) SEM of the basilar papilla in wildtype (D) and talpid3 (E) chickens. Arrows indicate cilia. Curved lines represent the base of stereocilia hair bundles. (F–K) Actin bundles identified by phalloidin (green) and centriolar localization identified by γ tubulin (red). (F′, G′) overview of wildtype and talpid3 basilar papilla, higher magnification in (F, G), red circles with line represent orientation of polarized actin bundles in basilar papilla; dashed red circles represent unpolarized actin bundles (Schematic L, M). (L′) Quantification of polarized haircells. (M′) Quantification of the angle of polarised hair cells. (H–K) Dashed white circles represent magnified images (Ji–Kii′). (Ji–Kii) White arrows indicate aligned centrosomes; blue arrows indicate unaligned centrosomes (Schematic N). (N′) Quantification of cells with aligned centrosomes. (O, P) Orientation based on placement of Golgi (TGN46, red) in comparison to actin indicating the leading edge (phalloidin, green) and nucleus (Dapi, blue, schematic in Q) in MEFs. Asterisks represent areas of higher magnification (not all represented at lower magnification). (Q′) Quantification of the angle of orientation of MEF cells in scratch assay. Scale Bars: A, B 5 mm; A′, A′′, B′, B′′ 1 mm; D, E 1 μm; F, G, H, I, J, K 20 μm; F′, G′ 100 nm; Ji, Ji′, Jii, Jii′, Ki, Ki′, Kii, Kii′ 10 μm; O, P 100 μm; Oi, Oii, Oiii, Oiv, Ov, Ovi, Pi, Pii, Piii, Piv, Pv, Pvi 25 μm.