Associate editor: G.J. Dusting
Therapeutic targeting of insulin-regulated aminopeptidase: Heads and tails?

https://doi.org/10.1016/j.pharmthera.2007.07.006Get rights and content

Abstract

Insulin-regulated aminopeptidase, IRAP, is an abundant protein that was initially cloned from a rat epididymal fat pad cDNA library as a marker protein for specialized vesicles containing the insulin-responsive glucose transporter GLUT4, wherein it is thought to participate in the tethering and trafficking of GLUT4 vesicles. The same protein was independently cloned from human placental cDNA library as oxytocinase and is proposed to have a primary role in the regulation of circulating oxytocin (OXY) during the later stages of pregnancy. More recently, IRAP was identified as the specific binding site for angiotensin IV, and we propose that it mediates the memory-enhancing effects of the peptide. This protein appears to have multiple physiological roles that are tissue- and domain-specific; thus the protein can be specifically targeted for treating different clinical conditions.

Introduction

Insulin-regulated aminopeptidase — IRAP; LNPEPEC; cystinyl aminopeptidase, oxytocinase, placental leucine aminopeptidase (P-LAP); EC 3.4.11.3.

The diverse functions of IRAP are reflected in its identification based on different physiological or biochemical properties by 3 different groups. In 1995, IRAP was purified and cloned by Keller et al. as a marker protein of GLUT4 vesicles (Keller et al., 1995). In the following year, it was cloned under the moniker oxytocinase as an aminopeptidase produced by the placenta and postulated to function as a degradative enzyme for oxytocin (OXY) as the name implies (Rogi et al., 1996). In 2001, IRAP was also identified as the angiotensin AT4 receptor, a binding site for the peptide angiotensin IV (Ang IV), characterized in a range of tissues, including the brain.

IRAP is a type II transmembrane protein of 1025 amino acid residues with 3 domains (Rogi et al., 1996, Laustsen et al., 1997, Rasmussen et al., 2000). The hydrophilic N-terminal intracellular domain is 110 amino acids in length. It contains motifs involved in endocytosis and trafficking (Keller et al., 1995) and interacts with a number of proteins. The hydrophobic transmembrane segment is 22 residues (amino acids 111–131) in length and is followed by an extracellular domain of 893 residues (amino acids 132–1025). The extracellular domain contains the catalytic site consisting of the GAMEN motif (amino acids 428–432) and the HEXXH(X)18E Zn2+-binding motif (amino acids 464–487), which identifies IRAP as an M1 aminopeptidase (Keller et al., 1995; Fig. 1).

Regulation of IRAP is of therapeutic potential based on the 3 known properties of the protein. This review will focus on the 2 functional domains of IRAP: (i) HEAD, which contains the catalytic site of IRAP — inhibition of the aminopeptidase activity in this domain has the potential to (a) facilitate memory and (b) induce labour; (ii) TAIL, the intracellular domain of IRAP — modulation of proteins that interact with this domain have the potential to enhance glucose uptake in insulin responsive cells providing a treatment for type 2 diabetes.

IRAP colocalizes with the insulin-responsive glucose transporter GLUT4 in insulin-responsive tissues, including skeletal muscle and adipose. In addition, IRAP is found in a number of other tissues, many of which do not express GLUT4, such as the adrenal gland, spleen, kidney medulla and placenta (reviewed in Chai et al., 2004). This broad tissue distribution supports the concept of a range of physiological roles for IRAP.

IRAP was originally identified as a marker protein of the specialized vesicles containing GLUT4; recycling with the glucose transporter within intracellular compartments in the basal state and accompanying the protein to the plasma membrane (PM) in response to insulin (reviewed in Bryant et al., 2002; Fig. 2). Mobilization of GLUT4 to the cell membrane by insulin facilitates a large influx of glucose into fat cells. The role of IRAP in insulin-responsive tissues is not fully defined, but it is proposed to play a role in the tethering of GLUT4 vesicles within the cell under basal conditions (see Section 3).

IRAP is also produced in the placenta during pregnancy, wherein it is present mainly in the apical membrane of syncytiotrophoblasts. The extracellular domain is proteolytically cleaved at the PM and is secreted into maternal serum as a soluble form (Yamahara et al., 2000). In contrast to the widespread tissue distribution of membrane-bound IRAP, the soluble form of the enzyme is only detected in serum during pregnancy. Serum levels of IRAP increase with advancing gestation until just before the onset of labor, before diminishing postpartum (Mizutani et al., 1996, Yamahara et al., 2000).

In the brain, IRAP is present in high concentrations in neurones in the CA1–CA3 regions of the hippocampus, frontal, prefrontal, insular and entorhinal cortices and basolateral amygdala — areas associated with cognition. However, the distribution of IRAP is not restricted to these sites in the brain as it is also found in all motor neurones and motor-associated regions, some hypothalamic, thalamic and sensory nuclei (Fernando et al., 2005). In neurones, IRAP is found predominantly in intracellular compartments, in vesicular-like structures. In some brain regions, IRAP exhibits a high degree of colocalisation with GLUT4, reminiscent of that observed in fat cells (Fernando et al., 2005; Fig. 3). However, unlike insulin-responsive cells, it remains unclear as to whether IRAP and/or GLUT4 translocation to the PM is responsive to a particular stimuli or simply constitutive.

Limited insights into the physiological importance of IRAP have been provided by a study on the IRAP knockout mouse (Keller et al., 2002). Although the IRAP null mice exhibited impaired insulin-stimulated glucose uptake in muscle and adipose cells, possibly due to the concurrent 40–85% decrease in GLUT4 levels in the different muscles and adipocytes, they maintained normal glucose homeostasis — fed and fasted blood glucose and insulin levels were indistinguishable from wild-type controls (Keller et al., 2002). The weights of adipose tissue and skeletal muscles were not significantly different from wild-type controls; however, their hearts were on average 20% larger — cardiomegaly has also been observed in the GLUT4 null mice. These mice displayed decreased GLUT4 levels of between 40% and 85% in the different muscles or in adipocytes (Keller et al., 2002). It is likely that tissue-specific IRAP knockout mice will yield more valuable information as has been the case for the studies on tissue specific GLUT4 knockout mice (Minokoshi et al., 2003).

Section snippets

Heads: extra cellular domain

IRAP belongs to the M1 family of aminopeptidases. Other members of this family include aminopeptidase A (APA), aminopeptidase B (APB), aminopeptidase N (APN) and leukotriene A4 hydrolase (LTA4H). The family is characterized by a catalytic site that consists 2 conserved sequence elements, the HEXXH(X)18E consensus Zn2+-binding motif and the GXMEN exopeptidase motif (Hooper, 1994). Residues immediately surrounding the catalytic site are highly conserved among the M1 members with amino acid

Tails: the intracellular domain

As stated above, IRAP is a type II integral membrane containing a 110-amino-acid cytoplasmic amino terminus. The cytoplasmic tail is highly conserved between species and contains 2 dileucine motifs, each with a preceding acidic cluster (Keller et al., 1995, Ross et al., 1996). The carboxy terminus of GLUT4 also contains a dileucine motif followed by an acidic cluster that are shown to be required for correct intracellular targeting of the transporter (Corvera et al., 1994, Garippa et al., 1996

References (96)

  • L. Eguez et al.

    Full intracellular retention of GLUT4 requires AS160 Rab GTPase activating protein

    Cell Metab

    (2005)
  • R.J. Garippa et al.

    The carboxyl terminus of GLUT4 contains a serine-leucine-leucine sequence that functions as a potent internalization motif in Chinese hamster ovary cells

    J Biol Chem

    (1996)
  • K.L. Grove et al.

    High salt intake differentially regulates kidney angiotensin IV AT4 receptors in Wistar-Kyoto and spontaneously hypertensive rats

    Life Sci

    (1999)
  • N.M. Hooper

    Families of zinc metalloproteases

    FEBS Lett

    (1994)
  • Y. Kakinuma et al.

    Anti-apoptotic action of angiotensin fragments to neuronal cells from angiotensinogen knock-out mice

    Neurosci Lett

    (1997)
  • T. Kamei et al.

    Interaction of Bnr1p with a novel Src homology 3 domain-containing Hof1p. Implication in cytokinesis in Saccharomyces cerevisiae

    J Biol Chem

    (1998)
  • S. Kane et al.

    A method to identify serine kinase substrates. Akt phosphorylates a novel adipocyte protein with a Rab GTPase-activating protein (GAP) domain

    J Biol Chem

    (2002)
  • S.R. Keller et al.

    Cloning and characterization of a novel insulin-regulated membrane aminopeptidase from Glut4 vesicles

    J Biol Chem

    (1995)
  • S.R. Keller et al.

    Mice deficient in the insulin-regulated membrane aminopeptidase show substantial decreases in glucose transporter GLUT4 levels but maintain normal glucose homeostasis

    J Biol Chem

    (2002)
  • H. Kozaki et al.

    Maternal serum placental leucine aminopeptidase (P-LAP)/oxytocinase and preterm delivery

    Int J Gynaecol Obstet

    (2001)
  • E.A. Kramar et al.

    Angiotensin II- and IV-induced changes in cerebral blood flow. Roles of AT1, AT2, and AT4 receptor subtypes

    Regul Pept

    (1997)
  • E.A. Kramar et al.

    Role of nitric oxide in angiotensin IV-induced increases in cerebral blood flow

    Regul Pept

    (1998)
  • E.A. Kramar et al.

    The effects of angiotensin IV analogs on long-term potentiation within the CA1 region of the hippocampus in vitro

    Brain Res

    (2001)
  • M. Larance et al.

    Characterization of the role of the Rab GTPase-activating protein AS160 in insulin-regulated GLUT4 trafficking

    J Biol Chem

    (2005)
  • P.G. Laustsen et al.

    The complete amino acid sequence of human placental oxytocinase

    Biochim Biophys Acta

    (1997)
  • J. Lee et al.

    Effect of intracerebroventricular injection of AT4 receptor ligands, Nle1-angiotensin IV and LVv-hemorphin 7, on spatial learning in rats

    Neuroscience

    (2004)
  • Y. Minokoshi et al.

    Tissue-specific ablation of the GLUT4 glucose transporter or the insulin receptor challenges assumptions about insulin action and glucose homeostasis

    J Biol Chem

    (2003)
  • I. Moeller et al.

    Angiotensin IV inhibits neurite outgrowth in cultured embryonic chicken sympathetic neurones

    Brain Res

    (1996)
  • N. Nakamura et al.

    The vesicle docking protein p115 binds GM130, a cis-Golgi matrix protein, in a mitotically regulated manner

    Cell

    (1997)
  • E.S. Pederson et al.

    Attenuation of scopolamine-induced spatial learning impairments by an angiotensin IV analog

    Regul Pept

    (1998)
  • E.S. Pederson et al.

    A role for the angiotensin AT4 receptor subtype in overcoming scopolamine-induced spatial memory deficits

    Regul Pept

    (2001)
  • G. Reed et al.

    A novel action of angiotensin peptides in inhibiting neurite outgrowth from isolated chick sympathetic neurons in culture

    Neurosci Lett

    (1996)
  • T. Rogi et al.

    Human placental leucine aminopeptidase/oxytocinase. A new member of type II membrane-spanning zinc metallopeptidase family

    J Biol Chem

    (1996)
  • S.A. Ross et al.

    Characterization of the insulin-regulated membrane aminopeptidase in 3T3-L1 adipocytes

    J Biol Chem

    (1996)
  • K. Sakamoto et al.

    Contraction regulation of Akt in rat skeletal muscle

    J Biol Chem

    (2002)
  • H. Sano et al.

    Insulin-stimulated phosphorylation of a Rab GTPase-activating protein regulates GLUT4 translocation

    J Biol Chem

    (2003)
  • M.F. Sardinia et al.

    AT4 receptor structure-binding relationship: N-terminal-modified angiotensin IV analogues

    Peptides

    (1994)
  • M. Sohda et al.

    Depletion of vesicle-tethering factor p115 causes mini-stacked Golgi fragments with delayed protein transport

    Biochem Biophys Res Commun

    (2005)
  • A. Subtil et al.

    Characterization of the insulin-regulated endocytic recycling mechanism in 3T3-L1 adipocytes using a novel reporter molecule

    J Biol Chem

    (2000)
  • T. Tominaga et al.

    Diaphanous-related formins bridge Rho GTPase and Src tyrosine kinase signaling

    Mol Cell

    (2000)
  • M. Tsujimoto et al.

    Identification of human placental leucine aminopeptidase as oxytocinase

    Arch Biochem Biophys

    (1992)
  • G. Vazeux et al.

    Identification of glutamate residues essential for catalytic activity and zinc coordination in aminopeptidase A

    J Biol Chem

    (1996)
  • S.B. Waters et al.

    The amino terminus of insulin-responsive aminopeptidase causes Glut4 translocation in 3T3-L1 adipocytes

    J Biol Chem

    (1997)
  • M.J. Wayner et al.

    Angiotensin IV enhances LTP in rat dentate gyrus in vivo

    Peptides

    (2001)
  • J.J. Westendorf

    The formin/diaphanous-related protein, FHOS, interacts with Rac1 and activates transcription from the serum response element

    J Biol Chem

    (2001)
  • J.J. Westendorf et al.

    Identification and characterization of a protein containing formin homology (FH1/FH2) domains

    Gene

    (1999)
  • J.W. Wright et al.

    Brain angiotensin receptor subtypes AT1, AT2, and AT4 and their functions

    Regul Pept

    (1995)
  • J.W. Wright et al.

    Angiotensin II(3-8) (ANG IV) hippocampal binding: potential role in the facilitation of memory

    Brain Res Bull

    (1993)
  • Cited by (0)

    View full text