Short sequence-paper Cloning, sequencing and expression in the seeds and radicles of two Lupinus albus conglutin Q genes 1 Alessio Scarafoni a ; *, Annalisa Di Cataldo a , Tatiana D. Vassilevskaia b , Evguenia P. Bekman b , Claudina Rodrigues-Pousada 2 ; b , Fabrizio Ceciliani c , Marcello Duranti a a Dipartimento di Scienze Molecolari Agroalimentari, Universita © degli Studi di Milano, Via Celoria 2, 20133 Milan, Italy b Instituto Gulbenkian de Cie ªncia, Apartado 14, 2781 Oeiras, Portugal c Istituto di Fisiologia Veterinaria e Biochimica, Universita © degli Studi di Milano, Via Celoria 10, 20133 Milan, Italy Received 29 August 2000; received in revised form 14 February 2001; accepted 4 April 2001 Abstract Two genes encoding conglutin Q have been isolated from a Lupinus albus genomic library and sequenced. The expression of conglutin Q was studied by partial amino acid sequencing of the mature seed protein and by nucleotide sequencing of reverse transcriptase^polymerase chain reaction products from various tissues during the plant life cycle. ß 2001 Elsevier Science B.V. All rights reserved. Keywords : Conglutin Q ; Leguminous plant; Germinating seed; Young root; Lupinus albus A number of major seed proteins have no storage role and in many cases no other evident biological function has been established so far. Conglutin Q (CQ) from Lupinus spp. is an example of these proteins [1]. CQ has previously been referred to as a storage protein, but due to several unusual features, this role has been reconsidered. Indeed, despite its high expression in the developing lupin seeds (up to 5% of the seed proteins) and deposition in the storage vacuoles [2], CQ dramatically di¡ers from the can- onical storage proteins because (i) it remains undegraded at germination [3,4]; (ii) it has been located in the extra- cellular spaces of germinating cotyledons [5] and in the young roots [6]; (iii) it is selectively secreted in high amounts upon heat treatment of lupin mature seeds [7]. CQ is a tetrameric protein, each monomer consisting of two disulphide-linked polypeptides of 30 and 17 kDa [8]; the large chain was shown to be glycosylated [9]. Signi¢- cant sequence homology of Lupinus angustifolius CQ (LangCQ) [10] has only been found with three proteins, all belonging to the plant kingdom. One is the 7S basic globulin (Bg7S) from soybean [11], which has been claimed to be part of the plant cellular signal transduction system due to its protein kinase activity [12], which is stimulated by leginsulin, a hormone-like peptide isolated from soybean radicles [13]. The second one is the extra- cellular dermal glycoprotein (EDGP) from carrot, which is expressed in response to wounding [14]. The third protein is a basic protein from cotton seeds showing a weak anti- fungal activity [15]. None of these activities have been identi¢ed for lupin CQ so far. Lupinus albus CQ (LalbCQ) has extensively been characterised at molecular level [16,17], but its nucleotide or amino acid sequences have as yet not been determined. In the present paper we de- scribe the cloning and sequencing of two CQ genes from L. albus and report the expression of the protein in develop- ing seeds and in the young roots of germinating seeds. These ¢ndings di¡er from those of Gayler and co-workers who detected only one version of the CQ gene in the L. angustifolius genome, which is almost exclusively expressed in developing seed cotyledons [18]. A genomic V-Dash library constructed from L. albus nuclear DNA [19] was screened by using a probe gener- ated by polymerase chain reaction (PCR) of genomic DNA isolated from L. albus, var. Multitalia, using the primers 5P-AAACCTCCTTGTTCTACCC-3P (sense) and 0167-4781 / 01 / $ ^ see front matter ß 2001 Elsevier Science B.V. All rights reserved. PII:S0167-4781(01)00225-1 * Corresponding author. Fax: +39-2-7063-3062; E-mail : alessio.scarafoni@unimi.it 1 EMBL accession numbers: AJ297490 and AJ297568. 2 Present address: ITQB, Stress and genomics, Apartado 127, 2781-901 Oeiras, Portugal. Biochimica et Biophysica Acta 1519 (2001) 147^151 www.bba-direct.com