Heat Shock Proteins and Modulation of Cellular Function
Role of AIP and its homologue the blindness-associated protein AIPL1 in regulating client protein nuclear translocation J. van der Spuy1 and M.E. Cheetham Division of Pathology, Institute of Ophthalmology, University College London, 11-43 Bath Street, London EC1V 9EL, U.K.
Abstract Mutations in the AIPL1 (aryl hydrocarbon receptor interacting protein-like 1) cause the blinding disease Leber’s congenital amaurosis. AIPL1 is a homologue of the AIP. AIP functions as part of a chaperone heterocomplex to facilitate signalling by the AhR and plays an important role in regulating the nuclear translocation of the receptor. We review the evidence for the role of AIP in protein translocation and compare the potential functions of AIPL1 in the translocation of its interacting partner the NEDD8 ultimate buster protein 1.
Introduction LCA (Leber’s congenital amaurosis) is inherited in an autosomal recessive manner and is characterized by severely impaired vision or blindness at birth. Mutations in AIPL1 (aryl hydrocarbon receptor interacting protein-like 1) cause up to 12% of LCA [1]. AIPL1 is a photoreceptor-specific protein and the expression of AIPL1 closely follows the spatiotemporal differentiation of rod and cone photoreceptors in the developing human retina [2]. In the adult human retina, however, AIPL1 is expressed only in the rod photoreceptors, suggesting a switch in AIPL1 function from growth to adulthood [3]. The function of AIPL1 is largely unknown but there are clues to its potential role in the retina from the many studies that have focused on the characterization of AIPL1 homologues.
AIPL1 homologues AIPL1 shares 49% identity and 69% similarity with the human AIP, also named the hepatitis B virus X-associated protein 2 or AhR (aryl hydrocarbon receptor)-activated protein 9 [4–6] (Figure 1). AIP in turn shares identity with the immunophilin co-chaperones FKBP (FK506binding protein) 51 and FKBP52 (Figure 1). AIP exists as a cytosolic ternary complex with the AhR, a ligandinducible transcription factor, which is a member of the basic helix–loop–helix Per-Arnt-Sim regulatory protein superfamily and a dimer of the molecular chaperone Hsp90 [4,6,7]. Similarly, the immunophilins cyp40, FKBP51 and FKBP52 are components of a cytosolic heterocomplex, including steroid hormone receptors and Hsp90. Three Key words: heat-shock protein, immunophilin, molecular chaperone, proteasome, retinal degeneration, tetratricopeptide repeat. Abbreviations used: AhR, aryl hydrocarbon receptor; AIP, AhR-interacting protein; AIPL1, AIPlike 1; Cul, cullin; FKBP, FK506-binding protein; LCA, Leber’s congenital amaurosis; NUB1, NEDD8 ultimate buster protein 1; TPR, tetratricopeptide repeat. 1 To whom correspondence should be addressed (email
[email protected]).
TPR (tetratricopeptide repeat) motifs are conserved among AIPL1, AIP and FKBP52/FKBP51 (Figure 1). The TPR is a degenerate motif that mediates protein–protein interactions [8]. The interaction of both AIP and FKBP52 with Hsp90 requires the TPR domain as well as a region C-terminal to the TPR domain [9–11]. AIP and FKBP52 bind to the C-terminus of Hsp90, which terminates in a conserved TPR domain recognition motif MEEVD [12–14].
Regulation of AhR translocation and transactivation by AIP (AhR-interacting protein) Functional studies of AIP have demonstrated that AIP enhances the ligand-induced transcriptional activity of the AhR [4,7,9,15,16]. In part, AIP is supposed to increase AhRmediated signal transduction by stabilizing the cytosolic levels of the AhR [10,16–18]. AIP can stabilize cytosolic AhR by protecting the AhR from ubiquitination and subsequent degradation mediated by the ubiquitin ligase protein, C-terminal Hsp70-interacting protein [16,19]. Furthermore, AIP is capable of modulating the intracellular distribution of the AhR. AIP mediates a distinct relocalization of ligand-free AhR to the cytoplasm and delays the nuclear translocation and accumulation of ligand-activated AhR (Figure 2) [19–21]. It has been suggested that AIP-mediated cytoplasmic retention of unactivated AhR could occur through the physical anchorage of the AhR to the actin cytoskeleton, although this finding has been disputed [21,22]. As an additional mechanism for the AIP-mediated cytoplasmic retention of the AhR, it has been shown that AIP can inhibit CRM-1mediated nucleocytoplasmic shuttling of the AhR and that cytoplasmic retention of the AhR is accomplished through the co-operative mechanisms of both AIP and CRM-1 (Figure 2) [21,22]. Alternatively, AIP may mediate AhR cytoplasmic retention by altering the conformation of the AhR nuclear localization signal, thereby hindering interaction C 2004
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Figure 1 Schematic representation of AIPL1, AIP and FKBP52 The pathological mutations in AIPL1 are shown. The primary structural features are labelled as follows: PP, polyproline-rich sequence; PPIase, peptidyl prolyl isomerase domain.
Figure 2 The function of AIP and AIPL1 (A) AIP regulates the nuclear translocation and transactivation of the AhR [10]. DRE, dioxin response element. (B) Similarly, AIPL1 modulates the nuclear translocation of its client protein, NUB1.
with the nuclear import receptor protein importin-β [21,23]. Therefore a number of mechanisms co-operate to mediate the cytoplasmic retention and translocation of the AhR by AIP.
ing of farnesylated proteins and it has been suggested that certain pathological mutations in AIPL1 compromise this activity [25]. The physiological significance of the interaction between AIPL1 and NUB1 is unknown; however, NUB1 interacts with the ubiquitin-like protein NEDD8 [26]. NEDD8 is conjugated specifically to members of the Cul (cullin) family in a manner analogous to ubiquitination and sentrinization [27]. The cullins are components of an SCF ubiquitin E3 ligase composed of Skp1, cullin, F-box protein and ROC1 [28]. The Cul-1 SCF complex catalyses the ubiquitination of inhibitory κBα, β-catenin, cyclin D proteins, p27 (KIP1) and p21 (CIP1/WAF1) [29–33], and NEDD8 conjugation is necessary for the ubiquitin ligase activity of the Cul-1 SCF complex [34,35]. The Cul-2 and Cul-3 SCF complexes target hypoxia-inducible factor-1α and cyclin E respectively for ubiquitination and proteasomal degradation [36,37]. Therefore NEDD8–cullin conjugation has been implicated in many important biological events, including cell signalling and cell-cycle regulation. NUB1 targets NEDD8 and NEDD8 conjugates for proteasomal degradation [38]. Recently, a splicing variant of NUB1 possessing a 14-aminoacid insertion in the C-terminus (NUB1L) was identified [39]. NUB1L has been shown to interact with NEDD8 as well as an additional ubiquitin-like protein FAT10, and to accelerate the proteasomal degradation of these ubiquitin-like proteins [39,40]. It has been suggested that the interaction of AIPL1 with NUB1 implicates AIPL1 in biological functions related to NEDD8 conjugation, including cell-growth and cell-cycle regulation, but the cellular role of NUB1 still remains to be fully determined.
The interacting partners of AIPL1 It is not known whether AIPL1, similar to AIP and FKBP52, is involved in the signalling of a nuclear receptor. Thus far, two independent studies using yeast two-hybrid screens have identified some interacting partners of AIPL1; NUB1 (NEDD8 ultimate buster protein 1) [24] and farnesylated proteins [25]. AIPL1 has been shown to assist the process C 2004
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AIPL1 modulates the subcellular distribution of NUB1 Recently, we examined the expression and localization of AIPL1 and NUB1 in the adult and developing human retina [2,3]. AIPL1 was predominantly cytoplasmic, whereas
Heat Shock Proteins and Modulation of Cellular Function
NUB1 was predominantly nuclear due to a functional nuclear localization signal near the C-terminus [2,26]. Therefore we tested the hypothesis that AIPL1 could modulate NUB1 nuclear translocation in a manner similar to the role of AIP with the AhR. Co-expression of AIPL1 and NUB1 resulted in a concentration-dependent modulation of NUB1 subcellular distribution towards the cytoplasm. Therefore there are parallels between the action of AIPL1 with NUB1 and AIP with the AhR (Figure 2). This effect appears to be specific for AIPL1, as AIP could not modulate the subcellular distribution of NUB1. These results suggest that AIPL1 may regulate the translocation of NUB1, thereby modulating downstream effects of NUB1 on cell signalling and cell growth.
Concluding remarks In conclusion, functional studies of AIPL1 to date would suggest a photoreceptor-specific function that is essential for retinal development and vision. Although few mechanistic details for AIPL1 function are known, there do appear to be similarities with the function of AIP since AIPL1 is capable of modulating the subcellular localization of its client protein NUB1. Further characterization of the AIPL1– NUB1 interaction will enhance our understanding of the physiological consequences of this association.
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