Angiotensin I-Converting Enzyme Gln1069Arg Mutation Impairs Trafficking to the Cell Surface Resulting in Selective Denaturation of the C-Domain

Abstract

BackgroundAngiotensin-converting enzyme (ACE; Kininase II; CD143) hydrolyzes small peptides such as angiotensin I, bradykinin, substance P, LH-RH and several others and thus plays a key role in blood pressure regulation and vascular remodeling. Complete absence of ACE in humans leads to renal tubular dysgenesis (RTD), a severe disorder of renal tubule development characterized by persistent fetal anuria and perinatal death.Methodology/Principal FindingsPatient with RTD in Lisbon, Portugal, maintained by peritoneal dialysis since birth, was found to have a homozygous substitution of Arg for Glu at position 1069 in the C-terminal domain of ACE (Q1069R) resulting in absence of plasma ACE activity; both parents and a brother who are heterozygous carriers of this mutation had exactly half-normal plasma ACE activity compared to healthy individuals. We hypothesized that the Q1069R substitution impaired ACE trafficking to the cell surface and led to accumulation of catalytically inactive ACE in the cell cytoplasm. CHO cells expressing wild-type (WT) vs. Q1069R-ACE demonstrated the mutant accumulates intracellularly and also that it is significantly degraded by intracellular proteases. Q1069R-ACE retained catalytic and immunological characteristics of WT-ACE N domain whereas it had 10–20% of the nativity of the WT-ACE C domain. A combination of chemical (sodium butyrate) or pharmacological (ACE inhibitor) chaperones with proteasome inhibitors (MG 132 or bortezomib) significantly restored trafficking of Q1069R-ACE to the cell surface and increased ACE activity in the cell culture media 4-fold.Conclusions/SignificanceHomozygous Q1069R substitution results in an ACE trafficking and processing defect which can be rescued, at least in cell culture, by a combination of chaperones and proteasome inhibitors. Further studies are required to determine whether similar treatment of individuals with this ACE mutation would provide therapeutic benefits such as concentration of primary urine.

Spectrum of mutations in the renin-angiotensin system genes in autosomal recessive renal tubular dysgenesis

“…All except those previously reported in four families [Danilov et al, 2010;Gribouval et al, 2005;Schreiber et al, 2010;Uematsu et al, 2009] were novel. Mutations were present in the homozygous state in 15 families, in the compound heterozygous state in 13, whereas only one heterozygous ACE mutation was identified in three families.…”

“…Mutations in the RAS genes are reported in 15 families [Bacchetta et al, 2007;Ben Amar et al, 2007;Danilov et al, 2010;Gribouval et al, 2005;Michaud et al, 2011;Schreiber et al, 2010;Uematsu et al, 2006Uematsu et al, , 2009Zingg-Schenk et al, 2008] and new sequence changes identified in our laboratory in 33 families are presented in Table 1. New mutations were identified by direct sequence analysis as described previously [Gribouval et al, 2005].…”

“…Since the initial publication, nearly 100 cases of sporadic or familial cases of RTD have been reported [Allanson et al, 1992;Ariel et al, 1995;Bacchetta et al, 2007;Ben Amar et al, 2007;Bernstein and Barajas, 1994;Danilov et al, 2010;Gribouval et al, 2005;Huang et al, 2003;Kriegsmann et al, 2000;Kumar et al, 1997;Kumral et al, 2003;Lacoste et al, 2006;MacMahon et al, 1990;Mantan et al, 2004;McFadden et al, 1997;Metzman et al, 1993;Michaud et al, 2011;Milunsky et al, 1997;Moldavsky, 2009;Querfeld et al, 1996;Russo et al, 1991;Schreiber et al, 2010;Schwartz et al, 1986;Swinford et al, 1989;Uematsu et al, 2006Uematsu et al, , 2009Voland et al, 1985;Zingg-Schenk et al, 2008]. Renal lesions are more diffuse than initially described: Henle loops and collecting ducts are collapsed whereas the muscular wall of medium-sized arteries and arterioles is thickened and disorganized [Ariel et al, 1995;Lacoste et al, 2006;Voland et al, 1985].…”

The iodide transport defect-causing mutation R124H: a δ-amino group at position 124 is critical for maturation and trafficking of the Na+/I− symporter (NIS)

“…This quality control mechanism ensures that only properly folded proteins are delivered to their final destinations, thereby retaining and degrading misfolded polypeptides without preventing the export of properly folded proteins. However, several human diseaserelated mutant proteins with varying degrees of activity are retained in the ER, such as Duox2, CFTR, aquaporin-2, angiotensin I-converting enzyme, the V2 vasopressin receptor, the sulfonylurea receptor, the HERG K + channel, the melanocortin-4 receptor and the luteinizing hormone G proteincoupled receptor (Danilov et al, 2010;Grasberger et al, 2007;Li et al, 1993;Meimaridou et al, 2011;Morello et al, 2000;Newton et al, 2011;Partridge et al, 2001;Tamarappoo and Verkman, 1998;Zhou et al, 1999). Most ITD-causing NIS mutants are, like WT NIS, targeted to the plasma membrane.…”

A Connexin50 Mutant, CX50fs, That Causes Cataracts Is Unstable, but Is Rescued by a Proteasomal Inhibitor

“…In fact, one drug of this class, bortezomib, is being used for treatment of some hematologic malignancies. Proteasomal inhibitors have also been reported to rescue targeting to the plasma membrane and function of some proteasomally degraded mutants associated with other diseases (45)(46)(47)(48). Other inhibitors of the proteasome that may have reduced secondary effects (49,50) are being developed, and they may eventually allow chronic application to prevent or ameliorate disease (like cataracts).…”