Neural Wiskott-Aldrich syndrome protein modulates Wnt signaling and is required for hair follicle cycling in mice
Anna Lyubimova, John J. Garber, Geeta Upadhyay, Andrey Sharov, Florentina Anastasoaie, Vijay Yajnik, George Cotsarelis, Gian Paolo Dotto, Vladimir Botchkarev, Scott B. Snapper
Anna Lyubimova, John J. Garber, Geeta Upadhyay, Andrey Sharov, Florentina Anastasoaie, Vijay Yajnik, George Cotsarelis, Gian Paolo Dotto, Vladimir Botchkarev, Scott B. Snapper
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Research Article Dermatology

Neural Wiskott-Aldrich syndrome protein modulates Wnt signaling and is required for hair follicle cycling in mice

Abstract

The Rho family GTPases Cdc42 and Rac1 are critical regulators of the actin cytoskeleton and are essential for skin and hair function. Wiskott-Aldrich syndrome family proteins act downstream of these GTPases, controlling actin assembly and cytoskeletal reorganization, but their role in epithelial cells has not been characterized in vivo. Here, we used a conditional knockout approach to assess the role of neural Wiskott-Aldrich syndrome protein (N-WASP), the ubiquitously expressed Wiskott-Aldrich syndrome–like (WASL) protein, in mouse skin. We found that N-WASP deficiency in mouse skin led to severe alopecia, epidermal hyperproliferation, and ulceration, without obvious effects on epidermal differentiation and wound healing. Further analysis revealed that the observed alopecia was likely the result of a progressive and ultimately nearly complete block in hair follicle (HF) cycling by 5 months of age. N-WASP deficiency also led to abnormal proliferation of skin progenitor cells, resulting in their depletion over time. Furthermore, N-WASP deficiency in vitro and in vivo correlated with decreased GSK-3β phosphorylation, decreased nuclear localization of β-catenin in follicular keratinocytes, and decreased Wnt-dependent transcription. Our results indicate a critical role for N-WASP in skin function and HF cycling and identify a link between N-WASP and Wnt signaling. We therefore propose that N-WASP acts as a positive regulator of β-catenin–dependent transcription, modulating differentiation of HF progenitor cells.

Authors

Anna Lyubimova, John J. Garber, Geeta Upadhyay, Andrey Sharov, Florentina Anastasoaie, Vijay Yajnik, George Cotsarelis, Gian Paolo Dotto, Vladimir Botchkarev, Scott B. Snapper

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Figure 2

Hair loss, ulceration, and epidermal hyperproliferation in N-WASP EKO mice.

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Hair loss, ulceration, and epidermal hyperproliferation in N-WASP EKO mi...
(A) WT and EKO littermate mice 9, 12, and 20 weeks old. Ulcers (arrows) on the chest and face of a 17-week-old EKO mouse are also shown. (B) Representative FACS analysis of primary keratinocytes. Keratinocytes were isolated from mice 2 hours after BrdU injection and analyzed by flow cytometry. Selected (gated) events represent the total percentage of BrdU+ keratinocytes in the representative experiment; average BrdU+ in EKO mice was 5.1% (n = 11) vs. WT 1.0% (n = 10) (P = 0.004). (C) Results of proliferation analysis of primary keratinocytes in culture. Cultured keratinocytes were labeled with BrdU for 30 minutes, fixed, stained with anti-BrdU antibody, and analyzed by flow cytometry. The results of 1 representative experiment (repeated 3 times) are shown; each bar represents mean value of 6 samples ± SD. (D) H&E-stained paraffin skin sections of WT and EKO mice demonstrate an expansion/hyperproliferation of HFs and interfollicular epidermis (upper panel, arrows). Frozen skin sections were stained with the antibody against the proliferation marker Ki-67 (green), and nuclei were stained with propidium iodide (PI, red). The increased epidermal proliferation was associated with an increase in Ki-67+ nuclei. Scale bar: 100 mm. (E) Normal distribution of early (keratin 1) and late (loricrin, filaggrin) markers of differentiation in hyperproliferative N-WASP–deficient epidermis. Frozen sections stained with antibodies against keratin 1, loricrin, and filaggrin (red). Nuclei were counterstained with Hoechst (blue). Scale bar: 40 mm.