Journal of Pathology J Pathol 2003; 201: 119–126. Published online 13 May 2003 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/path.1389 Original Paper Adducin expression in cutaneous and oral lesions: α - and β -adducin transcripts down-regulate with keratinocyte differentiation in stratified epithelia Sharn L Bowen, 1 Balvinder K Bloor, 1 Irene M Leigh 2 and Ahmad Waseem 1 * 1 Head and Neck Cancer Research Programme, GKT Dental Institute, King’s College London, London, UK 2 Cancer Research UK Skin Tumour Biology Unit, Centre for Cutaneous Research, The Royal London Hospital, London, UK *Correspondence to: Dr Ahmad Waseem, Head and Neck Cancer Research Programme, Floor 28, Guy’s Tower, London Bridge, London SE1 9RT, UK. E-mail: ahmed.waseem@kcl.ac.uk Received: 10 October 2002 Revised: 9 January 2003 Accepted: 7 March 2003 Abstract Adducin is a heterodimer of α- with β - or γ -subunits that regulates the assembly of the spectrin-based membrane skeleton in erythrocytes. Although adducin has been identified in various non-erythroid cells and tissues, it has been localized at intercellular junctions only in keratinocytes and epidermis. However, no data are available yet on the regulation of individual adducin genes in differentiating versus hyperproliferating keratinocytes. Due to the unavailability of mono-specific antibodies for individual adducins, this study has used RT-PCR and in situ hybridization to investigate the expression of α- and β -adducins in cultured cells and in stratified epithelia including cutaneous and oral lesions. Using RT- PCR, the α-transcripts were consistently expressed in all cell lines tested, as well as in normal interfollicular epidermis, whereas the β -transcripts were more variable and were strongly expressed in K562, A431, and primary keratinocytes. However, in normal skin, oral mucosa, and attached gingivae, the levels of α-transcripts closely paralleled those for the β -subunit. In most normal tissues, adducin expression was observed primarily in the proliferating compartments including the basal layer and lower suprabasal layers. Expression of both genes was also up-regulated in skin diseases characterized by increased cell proliferation and keratinocyte activation, such as basal cell carcinoma (BCC), squamous cell carcinoma (SCC), and psoriasis. It was observed that, in most cases, the expression of both α- and β -adducin was accompanied by increased expression of the proliferation marker Ki-67 and keratins K6 and K16. Differentiating keratinocytes in normal epithelia as well as in tumours appear to suppress the expression of adducin transcripts. The data suggest that the expression of adducin genes may be linked to cell proliferation and starts to down-regulate at the onset of differentiation. Copyright  2003 John Wiley & Sons, Ltd. Keywords: hyperproliferation; gene expression; in situ hybridisation; psoriasis; BCC; SCC; oral epithelia Introduction Adducin is a component of the two-dimensional net- work of proteins that is organized around spectrin and is referred to as the membrane skeleton. The struc- tural organization of the membrane skeleton has been studied most extensively in human erythrocytes (for a review see ref 1). The basic structure involves asso- ciation of spectrin with actin oligomers, stabilized by protein 4.1, into a two-dimensional lattice that is held to the membrane by the interaction of protein 4.1 and ankyrin to glycophorin and band 3, respectively [2,3]. Several proteins including dematin, adducin, and pro- tein 4.2 are associated with the membrane skeleton in a dynamic equilibrium regulated by Ca 2+ /calmodulin [4] and by phosphorylation [5,6]. Adducin is a heterodimer of α- and β -subunits, which were first identified in erythrocytes as a Ca 2+ /calmodulin-binding protein and the major sub- strate of PKC in erythrocytes [4,7]. A γ -adducin has also been described which forms heterodimers with α-adducin in the absence of the β -subunit [8]. Adducin appears to play a key role in the assembly of the membrane skeleton [4], regulated by phosphoryla- tion by several kinases including protein kinase C, protein kinase A, Rho kinase, and lyn, a Src tyro- sine kinase [9–11]. Mutations in adducin polypeptides have been implicated in hypertension in humans and rodents [12]. Non-erythroid adducin has been identi- fied in various tissues and cells, including the brain, kidney, epidermis, lens, MDCK cells, fibroblasts, and human keratinocytes [13–16]. Most of the studies on adducin expression have utilized polyclonal antibodies raised against erythroid or brain adducin complexes and no data have been published yet on the individual expression of α- and β -adducins in tissues. Furthermore, while α- and β -adducins are detected in epithelia by northern [13] and western blotting [15], there are no data on the relative levels of their transcripts in stratified epithelia. Copyright  2003 John Wiley & Sons, Ltd.