Human epidermis is a novel site of phospholipase B expression Eric Maury, a,b Marie-Claude Pr  evost, b Michel Nauze, b Daniel Redoul  es, a Roger Tarroux, a Marie Charv eron, a Jean-Pierre Salles, b Bertrand Perret, b Hugues Chap, b and Ama Gassama-Diagne b, * a Institut de recherche Pierre Fabre, CERPER/H^ otel Dieu Saint Jacques, F31052 Toulouse, France b Centre de Physiopathologie de Toulouse Purpan, Institut F ed eratif de Recherche en Immunologie Cellulaire et Mol eculaire, H^ opital Purpan, F31069 Toulouse, France Received 4 June 2002 Abstract Phospholipase B (PLB) is an enzyme that displays both phospholipase A 2 and lysophospholipase activities. Analysis of human epidermis homogenates indicated the presence of a 97kDa PLB protein, as well as a phospholipase A 2 activity, both being enriched in the soluble fraction. Immunolabelling and in situ hybridization experiments showed that this enzyme is expressed in the different layersofepidermiswithanaccumulationatthedermo-epidermisjunction.RT-PCRdataindicatedthatPLBisspecificallyexpressed innaturalandreconstructedepidermis.By3 0 -RACE-PCRandscreeningofhumangenomedatabases,weobtaineda3600bpcDNA codingforhumanPLBhighlyhomologoustoalreadydescribedintestinalbrushborderPLBs.Thesedataledustoconcludethatthe soluble PLB corresponds to a proteolytic cleavage of the membrane anchored protein. Altogether, our results provide the first characterization of human PLB which should play an important role in epidermal barrier function. Ó 2002 Elsevier Science (USA). All rights reserved. Keywords: Phospholipase B; Human; Epidermis; Keratinocyte; Molecular cloning Phospholipase B (PLB) is an enzyme that can remove both the sn-1 and sn-2 fatty acids of glycerophospholi- pids and thus displays phospholipase and lysophos- pholipaseactivities.SeveralPLBshavebeenidentifiedin various microorganisms [1], plants [2], and in the brush bordermembraneofmatureenterocytesfromguineapig [3], rat [4], and rabbit [5]. Moreover, PLB also displays lipase and retinyl ester hydrolase activities [6]. However, despite its broad substrate specificity, PLB preferentially hydrolyses the sn-2 positionofphospholipidsandcanbe viewed as a PLA 2 [7]. Molecular cloning of mammalian PLBs revealed a good conservation of this enzyme. The protein is anchored in the membrane by a single hy- drophobic segment, followed by a short C-terminal cy- toplasmic tail. The large extracellular domain is heavily glycosylated and consists of four tandem homologous domains. For rat PLB, in the repeat 2, the chymotryp- sin-like catalytic triad including Ser 404 , Asp 518 , and His 659 is essential for activity [8]. PLB is synthesized as a 170kDa proenzyme activated upon trypsin treatment to a protein of 140kDa [9]. However, a 97kDa active protein could be obtained upon papain treatment of intestinal brush border membrane [3,10]. PLB was first described in intestine [10], but was also identified in two additional epithelia sites: testis [11] and epididymis [9]. Although intestinal epithelium is com- posed of a monolayer of cells, differentiation between epidermis and intestine cells shares common features. The epidermis is composed of several cell layers. The deepest layer, located at the dermal–epidermal junction (basal layer), consists of undifferentiated keratinocytes, which proliferate. As cells migrate up through the epi- dermis, keratinocytes undergo terminal differentiation. This pattern is characterized by growth arrest and expression of the mature cytokeratins 1, 10 and profi- laggrin [12,13]. The corneocytes are terminally differen- tiated cells and constitute the outer layer of the epidermis, which provides a protective barrier [14]. Biochemical and Biophysical Research Communications 295 (2002) 362–369 www.academicpress.com BBRC * Corresponding author. Fax: +33-5-61-77-94-01. E-mail address: ama.gassama@toulouse.inserm.fr (A. Gassama- Diagne). 0006-291X/02/$ - see front matter Ó 2002 Elsevier Science (USA). All rights reserved. PII:S0006-291X(02)00657-5