Genomic Organization of the Human  Adducin Gene Lorena Citterio,* Tiziana Azzani,* Stefano Duga,† and Giuseppe Bianchi* *Division of Nephrology, Dialysis and Hypertension, San Raffaele Hospital, Milan, Italy; and †Veterinary Physiology and Biochemistry Institute, University of Milan, Milan, Italy Received October 16, 1999 We report the genomic structure of the human  adducin gene (ADD3). Adducin is a protein involved in cytoskeletal assembly and composed of – or – sub- units which share a high degree of homology between human and rat. Mutations in  subunit have been shown associated to both human and rat hyperten- sion. The human ADD3 gene spans over 20 kb and is composed of at least 13 introns and 14 exons covering the entire coding region. The exon size ranges from 81 bp to greater than 293 bp and the intron size from 111 bp to longer than 3.2 kb. We also demonstrate the presence of an alternative splicing event around exon 13, whose sequence, position, and expression is analo- gous in rat Add3 gene. Moreover, human ADD3 amino acid sequence presents 91.9% of identity compared to rat sequence. Characterization of human ADD3 gene provides an important tool for mutation analysis. © 1999 Academic Press Adducin is an heterodimeric cytoskeleton protein in- volved in the formation of actin-spectrin lactice, actin polimerization and signal transduction (1, 2). The pro- tein is composed of related but not identical subunits (,  and ); moreover  and  subunits show a very high homology between humans and rats (91.3% and 91.7% respectively). It has been postulated that adducin may partecipate in the regulation of blood pressure. In hypertensive rats of the Milan strain (MHS) a primary increase of renal tubular Na + reabsorption (3) is involved in the development of hypertension. In particular, the MHS strain shows an increased activity and expression of Na,K-pump units per cell compared to the normoten- sive MNS control (4). A missense mutation in  subunit (F316Y) is genetically associated to hypertension and this effect is modulated by mutations in  subunit (Q529R) (5) and in  subunit (Q572K) (L. Zagato per- sonal communication). Transfection studies have dem- onstrated that the – hypertensive variant of the protein modulates actin assembly and increases the surface expression and the activity of the Na,K-pump (6). In humans the -adducin locus is associated with and linked to essential hypertension (7–9). A missense mutation in this subunit (G460W) is associated with hypertension and affects the relationship between re- nal Na + excretion and blood pressure (10). Recent re- sults indicate a direct and specific interaction between adducin and Na,K-ATPase in vitro; both rat and hu- man adducin polymorphisms differently modulate re- nal Na,K-ATPase in vitro (11). ,  and -adducin genes (ADD1, ADD2, ADD3) are localized on different chromosomes in both humans and rats (12–14) and are differently expressed in a variety of tissues. -adducin is expressed in all tissues examined and may coexist as – and/or – het- erodimers depending on the expression levels of  and  genes in the different tissues (15). A transcript of 4 kb has been detected in human kidney mRNA using an adducin-like human cDNA clone as probe (16), confirm- ing the expression signal of  subunit and the absence of  signal in renal tissue (12). The expression of human ADD3 in the kidney sug- gests that ADD3 could have a role also in human hypertension. Therefore we set out to establish the exon-intron organization of the human ADD3 ORF for genomic DNA based mutation detection. MATERIALS AND METHODS Identification of human ADD3 and chromosomal mapping using database analysis. Human gamma adducin cDNA (complete coding sequence Accession Number U37122), obtained by database searches using locally maintained Genbank/EMBL database, was used to perform a BLASTN search against the STSs in the nonredundant Database of Genbank STS Division. Only one STS (Accession Num- ber G03246) showed a strong similarity to ADD3 cDNA. Further information were also reported: the linked markers and distance expressed in centiRadios from the top of chromosome linkage group, YAC clones issued by CEPH library. The nucleotide sequence data reported in this paper have been submitted to GenBank and have been assigned the Accession Num- bers Y14372 through Y14384. Biochemical and Biophysical Research Communications 266, 110 –114 (1999) Article ID bbrc.1999.1769, available online at http://www.idealibrary.com on 110 0006-291X/99 $30.00 Copyright © 1999 by Academic Press All rights of reproduction in any form reserved.