A Transposon-Based Analysis of Gene Mutations Related to Skin Cancer Development Rita M. Quintana, Adam J. Dupuy, Ana Bravo, M Llanos Casanova, Josefa.

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A Transposon-Based Analysis of Gene Mutations Related to Skin Cancer Development Rita M. Quintana, Adam J. Dupuy, Ana Bravo, M Llanos Casanova, Josefa P. Alameda, Angustias Page, Miguel Sánchez-Viera, Angel Ramírez, Manuel Navarro Journal of Investigative Dermatology Volume 133, Issue 1, Pages 239-248 (January 2013) DOI: 10.1038/jid.2012.245 Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 K5-SB11 is expressed in basal cells of skin and other stratified epithelia and can mobilize transposons in transgenic (Tg) mice. (a) Representation of the K5-SB11 transgene. (b) Northern blot analysis of RNA from back skin of Tg and wild-type (WT) mice. (c) Western blot analysis of total protein extracts from Tg and WT mice. (d) Immunohistochemical analysis of K5-SB11 transgene expression in adult back and tail skin. SB11 stains nuclei from hair follicles (arrowheads), bulge (asterisk), and basal epidermal cells (arrows). Bar=100μm. (e) Coexpression of SB11 (red) and CD34 (green) in hair follicle bulge cells (asterisk). (f) PCR analysis of transposition in T2Onc2 mice; the 225-bp band indicates transposition. (g) Southern blot analysis of transposition in DNA from skin epidermis. Ages of mice are indicated at the bottom of the panel. Lane 6: WT mouse. The filter was sequentially hybridized with probes for T2Onc2 (upper panel) and Thy1 (loading control, lower panel). Journal of Investigative Dermatology 2013 133, 239-248DOI: (10.1038/jid.2012.245) Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Increased malignancy in transposon/transposase tumors. (a) Squamous papilloma from control animals. (b–f) Squamous papillomas from transposon/transposase animals. (b) Papilloma with basal, pseudocarcinomatous cell proliferation and suprabasal acanthosis (hyperplasia). (c) Papilloma showing pseudocarcinomatous basal proliferation and intense spongiosis. (d) Papilloma showing basal atypia and diffuse basal membrane (arrowhead). (e) Papilloma with carcinoma in situ focus (arrow). (f) Papilloma with microcarcinoma focus (arrow). (g) Undifferentiated, anaplastic squamous cell carcinoma (SCC). (h–l) Basal cell carcinomas (BCCs) with areas of sebaceous differentiation (vacuolated clear cytoplasm of mature-like sebaceous cells in j, l) and foci of squamous differentiation (arrows in h, j). (k) BCC (arrow) adjacent to a SCC; i and j are two different regions of the same tumor and l is a magnification of k. Bar=50μm. Journal of Investigative Dermatology 2013 133, 239-248DOI: (10.1038/jid.2012.245) Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Immunohistochemical analysis of NSD1 expression in human skin and skin tumors. Control skin shows nuclear immunostaining of NSD1 (a, g), whereas basal cell carcinomas (BCCs) show reduced staining, with sporadic positive nuclei (b–e). Squamous cell carcinomas (SCCs) were positive (f) or negative (h). Panel g shows control (nontumoral) skin for the same histological sample as the SCC in h. Epidermis (Ep), BCC, and SCC locations are indicated. Bar=50μm. Journal of Investigative Dermatology 2013 133, 239-248DOI: (10.1038/jid.2012.245) Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions