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Yap1 deletion in neurons affects cerebellar morphology.

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posted on 2019-05-01, 17:25 authored by Katarzyna O. Rojek, Joanna Krzemień, Hubert Doleżyczek, Paweł M. Boguszewski, Leszek Kaczmarek, Witold Konopka, Marcin Rylski, Jacek Jaworski, Lars Holmgren, Tomasz J. Prószyński

(A) Strategy for generating Yap1 conditional knockout mice, showing the Yap1 flox allele (upper diagram) and recombined allele after Cre-mediated excision (lower diagram). (B) Lower Yap1 expression in the cerebellum of Yap1 fl/fl;Syn-Cre P30 mice analyzed by western blot. (C) Yap1 fl/fl;Syn-Cre mice exhibited a smaller cerebellum compared with control sex-matched littermates (Yap1 fl/fl). (D) Quantitative analysis of cerebellar weight in Yap1 fl/fl (n = 10) and Yap1 fl/fl;Syn-Cre (n = 4) P30 mice. p = 0.0178. Measurements were normalized to whole-brain weight. (E) Sagittal cerebellar sections of Yap1 fl/fl and Yap1 fl/fl;Syn-Cre P30 mice immunolabeled for calbindin (green) to visualize Purkinje cells and the molecular layer and NeuN (red) to visualize the granule cell layer. Numbers indicate individual lobes. (F) Quantification of Purkinje cells in sagittal sections of Yap1 fl/fl mice (n = 4 mice and at least three sections per mouse) and Yap1 fl/fl;Syn-Cre mice (n = 4 mice and at least three sections per mouse). p = 0.1782. (G) Decrease in cerebellar molecular layer thickness in Yap1 fl/fl;Syn-Cre mice visualized with calbindin; lobe III down the preculminate fissure at P30. (H-J) Thickness of the molecular (H), granular (I), and Purkinje (J) cell layers in the cerebellum of Yap1 fl/fl (n = 4 mice and at least three sections per mouse) and Yap1 fl/fl;Syn-Cre (n = 4 mice and at least three sections per mouse) P30 mice measured in lobe III down the preculminate fissure. p = 0.0128, p = 0.4679, and p = 0.1885. (K, L) Granule cell distribution (K) and quantification (L) in the molecular layer in P30 Yap1 fl/fl mice (n = 4 mice and at least three sections per mouse) and Yap1 fl/fl;Syn-Cre mice (n = 4 mice and at least three sections per mouse) in the cerebellum, shown as a percentage of NeuN-positive cells per 20,000 μm2. p = 0.0010. Numerical values that underlie the graphs are shown in S1 Data. Statistical significance was analyzed using two-tailed unpaired t test. *p < 0.05, **p < 0.01. Bars represent the mean ± SD. Scale bar = 5 mm in C, 1,000 μm in E, and 20 μm in G and K. ns, not significant; P, postnatal day; SD, standard deviation; Yap1, Yes-associated protein 1.

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