Evolution of aspartyl proteases by gene duplication: the mouse renin gene is organized in two homologous clusters of four exons.

Abstract

Overlapping recombinant clones that appear to encompass the entire renin gene, named Ren 1, have been isolated from a library of BALB/c mouse genomic DNA fragments. Based on restriction endonuclease mapping and DNA sequence analysis, Ren 1 spans 9.6 kb and contains nine exons interrupted by eight intervening sequences of highly variable size. The first exon, encoding the signal peptide of preprorenin, is separated from the eight following exons by a 3-kb intron. These eight exons are organized into two clusters of four separated by a 2-kb intron. DNA stretches encoding the aspartyl residues, which are part of the active site of renin, are located at homologous positions in both clusters. Our results show that aspartyl protease genes have arisen by duplication and fusion of an ancestral gene containing five exons. The estimated date of the duplication event of the mouse renin genes Ren 1 and Ren 2 is discussed.Key words: kidney renin/protein glycosylation/rate of divergence/exon-intron boundaries Introduction Aspartyl protease represents one of the protein families most completely characterized by primary amino acid sequence and crystallographic studies. The aspartyl proteases active site contains two aspartate residues localized in two short amino acid stretches with sequence homology to one another, and separated by a distance of about half of the polypeptide chain (Sepulveda et al., 1975). Aspartyl proteases such as penicellopepsin have a bilobal structure in which the two lobes are related by a 2-fold symmetry axis (Hsu et al., 1977). These observations have led to a hypothesis that aspartyl protease genes have evolved by duplication and fusion of an ancestral gene coding for a 15 000 -20 000 dalton polypeptide having a fold similar to that of one lobe of pepsin (Tang et al., 1978).In contrast to the other aspartyl proteases, renin has an optimal activity at neutral pH and a substrate specificity restricted to the cleavage of the prohormone angiotensinogen. The primary source of renin is the kidney where its concentration is extremely low. However, in some mouse strains, high levels of renin activity are found in the submaxillary gland (SMG) of males (Wilson et al., 1977). The amino acid sequence of the SMG renin has been recently reported (Panthier et al., 1982a;Misono et al., 1982). Comparison of the amino acid sequence of renin and pepsin shows -420Wo homology. The two most homologous regions are those surrounding the active sites aspartates. These results strongly suggest that renin and pepsin genes derive from a common ancestor.

Identification of Bona Fide Alternative Renin Transcripts Expressed Along Cortical Tubules and Potential Roles in Promoting Insulin Resistance In Vivo Without Significant Plasma Renin Activity Elevation

“…In addition, PCR products excised from agarose gels were also sequenced directly. Sequences of gene-specific primers such as gene-specific primer 1 and gene-specific primer 2 were generated as follows according to the sequence of both Ren1C and Ren2 as demonstrated by Holm et al 14 : gene-specific primer 1 (Ex2-2) 5′ CAG GTA GTT GGT GAG GAC CAC G 3′(213-234) and gene-specific primer 2 (Ex2) 5′ GG/AT AAG ATT/C G/AGT CAA GGA AGG/A 3′ (187-207), respectively.…”

Biochemical, Structural and Pathophysiological Aspects of Prorenin and (Pro)renin Receptor

“…Prorenin is also synthesized in many other tissues like adrenal glands (Ganten et al, 1974(Ganten et al, , 1976Ho and Vinson, 1998), zona glomerulosa (Doi et al, 1984;Brecher et al, 1989), eye, Müller cells, mast cells (Krop et al, 2008), ovarian follicular fluid (Glorioso et al, 1986), and theca cells (Do et al, 1988), uterus (Derkx et al, 1987;Itskovitz et al, 1987), myometrium/decidual cells (Shaw et al, 1989), placenta (Lenz et al, 1991), chorionic cells, testis and leydig cells (Sealey et al, 1988). The submandibular gland in some mice strains produces a large amount of renin, which is a product of the Ren-2 renin gene distinct from the renal renin gene, Ren-1 (Cohen et al, 1972;Wilson et al, 1981;Holm et al, 1984) and this action is mediated by prorenin converting enzyme present in submandibular gland of the same mice strains (Kim et al, 1991). Prorenin, in the juxtaglomerular cells of the kidney, is converted to mature renin by the limited endoproteolysis after paired basic residues, Lys-Arg to remove the 43-amino acid residues containing prosegment sequence.…”

Ren-1 and Ren-2 loci are expressed in mouse kidney.

“…For example, suitable sequence differences exist between the Ren-J(C57) and Ren-J(DBA) alleles (6,18) to permit direct quantitation of their respective mRNAs in kidney. If the assays are used in conjunction with appropriate genetic crosses, they should be useful in genetically defining allele-specific differences.…”

“…All strains of inbred mice carry the Ren-J structural gene, which encodes the aspartyl protease renin. Some strains carry an additional renin gene, designated Ren-2, which apparently arose from a relatively recent gene duplication event (1)(2)(3)(4)6). The presence of the Ren-2 gene correlates with the highsubmaxillary gland (SMG) renin phenotype (1-3, 7); strains that carry both Ren-J and Ren-2 exhibit SMG renin levels that are elevated 100-fold as compared with strains which carry only A considerable body of evidence supports the hypothesis that Ren-2 is the predominantly expressed renin gene in the SMG of two-gene strains.…”