In Vivo Role of Vitamin D Receptor Signaling in UVB-Induced DNA Damage and Melanocyte Homeostasis Sharmeen Chagani, Sergiy Kyryachenko, Yoko Yamamoto,

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In Vivo Role of Vitamin D Receptor Signaling in UVB-Induced DNA Damage and Melanocyte Homeostasis Sharmeen Chagani, Sergiy Kyryachenko, Yoko Yamamoto, Shigeaki Kato, Gitali Ganguli-Indra, Arup K. Indra Journal of Investigative Dermatology Volume 136, Issue 10, Pages 2108-2111 (October 2016) DOI: 10.1016/j.jid.2016.06.004 Copyright © 2016 The Authors Terms and Conditions

Figure 1 Melanocyte-specific VDR ablation reduces number of differentiated melanocytes after UVB exposure. (a) Targeting of VDR allele. Flags indicate LoxP: P1/P2 and P1/P3 primers for L2 and L– allele, respectively. (b) PCR genotyping of Cre transgene and VDR allele. (c) Anti-TYRP1 (green) and anti-VDR (red) co-labeling of skin, counterstained with DAPI (blue). Dotted line indicates epidermal-dermal junction. Scale bar = 100 μm. (d) Scheme for acute UVB treatment. (e) Anti-TYRP1 (green) labeling of skin for TYRP1+ melanocytes. Dotted line indicates epidermal-dermal junction. Scale bar = 20 μm. (f) Graph represents TYRP1+ melanocytes per field. (g) FM staining of differentiated melanocytes. Black arrows indicate Fontana Masson positive differentiated and pigmented melanocytes. Scale bar = 20 μm. (h) Graph indicates FM+ melanocytes per field. ∗P ≤ 0.05, ∗∗P ≤ 0.01. n = 5 per group. Data represent mean ± standard error of the mean. B, BamHI; D, dermis; E (in a), EcoRI; E (in c, e, and g), epidermis; E2, exon 2; E3, exon 3; E4, exon 4; FM, Fontana-Masson; h, hours; mins, minutes; TYRP1, tyrosinase-related protein 1; WT, wild type. Journal of Investigative Dermatology 2016 136, 2108-2111DOI: (10.1016/j.jid.2016.06.004) Copyright © 2016 The Authors Terms and Conditions

Figure 2 Melanocyte-specific VDR ablation reduces precursor, proliferating, and apoptotic melanocytes and alters DNA damage after UVB exposure. (a) Anti-TYRP2 (red) and anti-PAX3 (green) co-labeling of PAX3+ melanocytes. (b) Bar graph represents PAX3+/TYRP2+ melanocytes per field. (c) Anti-PCNA (red) and anti-TYRP1 (green) co-labeling of TYRP1+ proliferating melanocytes. (d) Graph indicates percentage of proliferating melanocytes. (e) Anti-TYRP1 (green) and anti-CPD (red) co-labeling of CPD+ melanocytes. (f) Graph represents percentage of CPD+/TYRP1+ melanocytes. (g) TUNEL (green) and TYRP1 (red) co-staining for apoptotic melanocytes. (h) Graph shows percentage of TUNEL+/TYRP1+ melanocytes. DAPI counterstaining (blue) in a, c, e, and g. Dotted lines in a, c, e, and g indicate epidermal-dermal junction. Scale bar = 20 μm. ∗P ≤ 0.05, ∗∗P ≤ 0.01, ∗∗∗P ≤ 0.001. n = 5/group. Data represent mean ± standard error of the mean. CPD, cyclobutane pyrimidine dimer; D, dermis; E, epidermis; h, hours; mins, minutes; PCNA, proliferating cell nuclear antigen. Journal of Investigative Dermatology 2016 136, 2108-2111DOI: (10.1016/j.jid.2016.06.004) Copyright © 2016 The Authors Terms and Conditions