Expression of T-Cadherin in Basal Keratinocytes of Skin Shuxia Zhou, Norihisa Matsuyoshi, Yoshiki Miyachi  Journal of Investigative Dermatology  Volume 118, Issue 6, Pages 1080-1084 (June 2002) DOI: 10.1046/j.1523-1747.2002.01795.x Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Keratinocytes expressed mRNA of T-cadherin. RNA extracted from Pam212 cells, NIH3T3 cells, and B16 cells was reverse-transcribed to cDNA using superscriptII and oligo-dT primer. cDNA was amplified by PCR using specific primers for each kind of cadherin molecule and β-actin. Pam212 cells expressed T-cadherin besides E- and P-cadherin, whereas NIH3T3 and B16 cells expressed N-cadherin. Journal of Investigative Dermatology 2002 118, 1080-1084DOI: (10.1046/j.1523-1747.2002.01795.x) Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Differential distribution pattern of T-cadherin and the classical cadherins, E- and P-cadherin, in mouse keratinocytes. Pam212 cells were immunoreacted with the rabbit anti-T-cadherin antibody (a, d), rat antimouse E-cadherin, ECCD2 (b, e), and rat antimouse P-cadherin, PCD-1 (c, f). Rabbit anti-T-cadherin antibody was detected by Oregon green-conjugated goat antirabbit IgG; ECCD2 and PCD1 were detected by FITC-conjugated rabbit antirat IgG. Cell nuclei were stained with propidium iodine. Parts (a)–(f) are single stainings of T- (a, d), E- (b, e), and P-cadherin (c, f). Parts (g)–(i) are double stainings of T-cadherin and E-cadherin: (g) T-cadherin; (h) E-cadherin; (i) overlay of T- and E-cadherin. Part (j) is a Z-section of 3D scanning on a single cell immunostained with T-cadherin. Parts (k)–(m) are XZ-sections of 3D scanning on a cluster of cells immunostained with (k) T-, (l) E-, and (m) P-cadherin. T-cadherin (a, d, g, j, k) was diffusely distributed on the cell surface, whereas E-cadherin (b, e, h, l) and P-cadherin (c, f, m) were concentrated on the cell–cell border. Scale bar: 20 µm. Journal of Investigative Dermatology 2002 118, 1080-1084DOI: (10.1046/j.1523-1747.2002.01795.x) Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 T-cadherin is expressed on the basal cell layer of mouse skin.(a) The frozen section of mouse back skin was immunostained with rabbit anti-T-cadherin antibody, and then incubated with Cy3-conjugated antirabbit IgG secondary antibody. Only the basal cell layer was immunostained with T-cadherin. Part (b) is the high magnification of (a). Arrows show the apical and lateral sides of basal cells; arrowheads show the basal side of the basal cells. T-cadherin was expressed on both cell–cell contact regions and the basal side of basal cells. In double immunofluorescence staining (c–h), anti-T-cadherin antibody immunolabeling (d, g) was detected by Oregon green-conjugated goat antirabbit IgG secondary antibody (green), ECCD2 (c), or PCD-1. (f) monoclonal antibody immunolabeling was detected by Cy3-conjugated goat antirat IgG secondary antibody (red). The yellow color show the colocalized regions (e, h). Scale bar: (a, b) 20 µm, (c–h) 40 µm. Journal of Investigative Dermatology 2002 118, 1080-1084DOI: (10.1046/j.1523-1747.2002.01795.x) Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Human keratinocytes and skin also expressed T-cadherin.(a) Immunostaining of T-cadherin on DJM-1 cells. DJM-1 cells were firstly incubated with rabbit anti-T-cadherin antibody, and then reacted with Cy3 labeled goat antirabbit secondary antibody. Nuclei were stained with propidium iodine. Scale bar: 2 µm. (b) Western blot of T- and E-cadherin on DJM-1 cell lysate. The cell lysate was separated by SDS-PAGE, and then transferred to nitrocellulose membrane. The membrane was incubated with rabbit antihuman T-cadherin or mouse antihuman E-cadherin antibody, and then incubated with HRP-conjugated second antibody. Signals were detected by ECL system. Lane 1 is T-cadherin; lane 2 is E-cadherin. The molecular weight marker was indicated at the left side. (c) Immunostaining of T-cadherin on normal human skin. The frozen section of normal human skin was immunostained with rabbit anti-T-cadherin antibody, and then incubated with Cy3-conjugated antirabbit IgG secondary antibody. Only the basal cell layer expressed T-cadherin. Scale bar: 50 µm. Journal of Investigative Dermatology 2002 118, 1080-1084DOI: (10.1046/j.1523-1747.2002.01795.x) Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions