The Lipocalin Apolipoprotein D Functional Portrait: A Systematic Review

Abstract

Apolipoprotein D is a chordate gene early originated in the Lipocalin protein family. Among other features, regulation of its expression in a wide variety of disease conditions in humans, as apparently unrelated as neurodegeneration or breast cancer, have called for attention on this gene. Also, its presence in different tissues, from blood to brain, and different subcellular locations, from HDL lipoparticles to the interior of lysosomes or the surface of extracellular vesicles, poses an interesting challenge in deciphering its physiological function: Is ApoD a moonlighting protein, serving different roles in different cellular compartments, tissues, or organisms? Or does it have a unique biochemical mechanism of action that accounts for such apparently diverse roles in different physiological situations? To answer these questions, we have performed a systematic review of all primary publications where ApoD properties have been investigated in chordates. We conclude that ApoD ligand binding in the Lipocalin pocket, combined with an antioxidant activity performed at the rim of the pocket are properties sufficient to explain ApoD association with different lipid-based structures, where its physiological function is better described as lipid-management than by long-range lipid-transport. Controlling the redox state of these lipid structures in particular subcellular locations or extracellular structures, ApoD is able to modulate an enormous array of apparently diverse processes in the organism, both in health and disease. The new picture emerging from these data should help to put the physiological role of ApoD in new contexts and to inspire well-focused future research.

Keywords: ApoD; extracellular vesicles; lipid peroxidation; lipoprotein particles; lysosome; membrane management; oxidative stress; protein physiology.

Figure 1

ApoD literature search and inclusion criteria. (A) Yearly timeline of articles recovered by PubMed using the search query designed for our review. (B) PRISMA flow diagram for record inclusion in our review.

Figure 2

ApoD protein sequence and gene features. (A) Multiple sequence analysis (MSA) of the mature amino acid sequence of selected vertebrate species recovered from GenBank (Apla, Anas platyrhynchos_EOB05196.1; Caur, Cathartes aura_KFP53002.1; Ggal, Gallus gallus_NP001011692.1; Pcri, Pelecanus crispus_KFQ60274.1; Pcrs, Podiceps cristatus_KFZ69168.1; Prub, Phoenicopterus ruber_KFQ85568.1; Bmut, Bos mutus_ELR54927.1; Cela, Cervus elaphus_ABB77207.1; Chir, Capra hircus_XP005675150.1; Oari, Ovis aries_XP004003075.1; Sscr, Sus scrofa_XP001926098.2; Fcat, Felis catus_XP006936237.1; Umar, Ursus marinus_XP008706566.1; Oorc, Orcinus orca_XP004278821.1; Mbra, Myotis brandtii_EPQ12038.1; Pale, Pteropus Alecto_XP006906222.1; Ocun, Oryctolagus cuniculus_ NP001075727.1; Hsap, Hoo sapiens_ NP001638.1; Ptro, Pan troglodites_XP516965.1; Fdam, Fukomys damarensis_KFO33128.1; Mmus, Mus musculus_CAA57974.1; Rnor, Rattus norvegicus_NP036909.1). Asterisks represent identical residues in all sequences, and dots/double dots point to similar residues. α-helices and β-strands are shown on top of the MSA, based on the solved tertiary structure of human ApoD. Colored residues are: four conserved cysteines involved in intramolecular disulfide bonds (pink), the human-specific unpaired cysteine (purple), conserved tryptophan residue in the ligand binding pocket (green), two glycosylated Asn residues (yellow), the antioxidant Met residue (blue), and residues in the hydrophobic surface patches at the rim of the binding pocket (orange). (B) Schematic representation of the chromosomal location of ApoD gene in human, mouse and chicken genomes. (C) Schematic representation of a consensus gene architecture for chordate ApoD, with four coding sequence (CDS)-containing exons and several 5′-UTR exons.

Figure 3

Molecular features of ApoD. (A,B) Graycolored space-filled views of the human ApoD tertiary structure (modelled from PDB ID:2HZQ) showing charged residues in A (positive, red; negative, blue) and hydrophobic residues in B (orange). Side view of the β-barrel (left image; curved arrows point to the pocket entrance) and top view (right image) looking into the hydrophobic pocket (asterisk). (C–E) Human ApoD (PDB ID:2HZQ) side and top views with highlighted relevant residues. Colored residues in (C) are the antioxidant Met93 (blue); the human-specific unpaired Cys116 (purple); the conserved ligand binding pocket Trp127 (green); and the two glycosylated Asn45/Asn78 (yellow). Pink-colored residues in D are the four cysteines forming two intramolecular disulfide bonds. Orange-colored residues in E are those forming three hydrophobic loops around the pocket entrance. (F) Space-filled view of human ApoD with reported oligosaccharides linked to Asn45 and Asn78, as modelled by GlyProt (see Methods). (G) Cartoon representations of human ApoD dimers formed by hydrophobic patches (orange) or by intermolecular Cys116 disulfide bonds (pink). Variations of the particular configuration shown are possible. Dashed lines delineate the ligand pocket. (H) Representation of the best supported tetrameric structure of human ApoD found in BCF. Asterisks mark the ligand pocket accessible in all subunits (two facing back). Oligosaccharides shown in red. (I) Cartoon illustration of a side view of human ApoD with AA (red) and HpETE (blue) positioned into the hydrophobic pocket (marked by a dashed line) and interacting with the Met93-containing hydrophobic patch respectively. (J,K) Cartoon illustration of human ApoD interacting with higher-order lipid structures via the hydrophobic patches at rim of the pocket; (J) HDL particle; (K) Unilamellar vesicle (liposome).

Figure 4

Publications on ApoD expression and disease relationships. (A) Distribution of publications describing ApoD mRNA expression or protein presence in vivo, distributed by physiological systems and in cell cultures (primary cells or cell lines). (B) Publications with information on ApoD relationship to disease (expression changes triggered by disease or treatments, or association of ApoD gene variants with disease).

Figure 5

Factors and pathways regulating the expression of ApoD. (A) Diverse stimuli regulate ApoD expression in a variety of cells and physiological conditions. (B) Summary of upstream regulatory pathways regulating ApoD expression where elements of the signaling cascade have been identified.

Figure 6

Schematic representation of ApoD subcellular traffic. A model of an ApoD-expressing cell is represented. Canonical exocytosis through the RER-Golgi pathway generates the mature, glycosylated (red dots) protein. The tetrameric form identified in the breast cyst fluid is represented as the format detected in extracellular fluids. Once at the plasma membrane, ApoD can be endocytosed (by non-expressing cells as well) and targeted to lysosomes and autophagolysosomes. When endolysosomes develop into multivesicular bodies, ApoD would be carried on the outer surface of exosomes. Finally, ApoD can be transferred to HDL during their biogenesis, or during their lipid-efflux activity (upon HDL-receptor interaction).

Figure 7

ApoD physiology summary. (A) Schematic view of the lipid-managing biochemical function of ApoD. The lipid structure depicted can equally represent the surface of a lipoprotein particle, extracellular vesicle or cellular membrane. ApoD antioxidant activity can be maintained by redox cycling, requiring a reductase activity, or the cycle can terminate by oligomerization of oxidized ApoD. (B) Summary of global tissue function of ApoD, where it contributes to the turnover and maintenance of tissues and organs. This equilibrium is reached after developmental processes in which ApoD is also involved, and switches to a different state upon disease, injury or physiological aging.