LRP5 and LRP6 in development and disease

Abstract

Low-density lipoprotein-related receptors 5 and 6 (LRP5/6) are highly homologous proteins with key functions in canonical Wnt signaling. Alterations in the genes encoding these receptors or their interacting proteins are linked to human diseases, and as such they have been a major focus of drug development efforts to treat several human conditions including osteoporosis, cancer, and metabolic disease. Here, we discuss the links between alterations in LRP5/6 and disease, proteins that interact with them, and insights gained into their function from mouse models. We also highlight current drug development related to LRP5/6 as well as how the recent elucidation of their crystal structures may allow further refinement of our ability to target them for therapeutic benefit.

Figure 1. Function and Regulation of Lrp5 and Lrp6 in β-catenin Signaling

In the absence of an upstream signal, Lrp5/6 and Frizzled are inactive (1). This results in the cytoplasmic pool of β-catenin being recruited into a multi-protein destruction complex that includes Axin, APC, Casein kinase 1a (CK1), and Glycogen synthase kinase 3 (GSK3) [99]. This destruction complex facilitates the GSK3-dependent phosphorylation of β-catenin and its subsequent ubiquitinylation (Ub) by the E3 ligase, β-TrCP, which targets β-catenin for degradation via the proteasome (3). Several mechanisms have been identified by which Lrp5/6 availability for signaling is regulated. First, several secreted inhibitors can bind directly to Lrp5/6 and prevent activation by Wnt ligands. These include Dkk1, Sost, and Wise (2). In addition, the E3 ubiquitin transmembrane ubiquitin ligase ZNRF3 (and the related RNF43) normally decreases the stability of the Lrp5/6:Frizzled complex by targeting the complex for degradation (4). However, in the presence of R-spondin proteins, a complex is formed between ZNRF3, R-spodin (Rspo), and the seven transmembrane receptor, LGR4, that results in the membrane clearance of ZNRF3 (4), which consequently leads to increased availability of Lrp5/6:Frizzled receptor and potentiated signaling [4]. Activation of Lrp5/6 can occur by several mechanisms. The best known occurs when a Wnt ligand engages a Frizzled receptor, and Lrp5 or Lrp6 (5) [2]. This results in the phosphorylation of the carboxyl terminus of Lrp5/6, creating a binding site for Axin. The formation of this new complex (“inhibited destruction complex”) results in β-catenin remaining in this complex because of failure to recruit β-TrCP for ubiquitinylation, although GSK3-mediated phosphorylation still occurs. Eventually, this results in the saturation of inhibited destruction complex with β-catenin (8). The complex is no longer available to bind and target newly synthesized β-catenin for degradation. This leads to increased β-catenin levels in the cytoplasm and nucleus where it is now available to complex with members of the LEF/TCF family to regulate transcription of target genes (9). Norrin can also activate β-catenin signaling in a Frizzled-4 and Lrp5-dependent manner, although no evidence for direct binding of Norrin to Lrp5 has been reported [100]. Finally, recent evidence [10] has found that Complement C1q can bind to Frizzled and activate C1s to cleave Lrp5/6, creating a truncated version of this protein that is constitutively active (7).

Figure 2. LRP5 and LRP6 are Highly Homologous Proteins with Similarly Arranged Structural Motifs

A schematic representation of human LRP6 is depicted with specific domains indicated by the color codes shown. The boundaries between each of the domains are indicated by the numbers listed at the top (relative to the 1613 amino acid full length protein). The levels of homology and identity between human LRP6 and human LRP5 are shown.

Figure 3. Structural Insights into the Function and Regulation of Lrp5/6

(A) Schematic representation created with the Pymol program of the ECD of LRP6 with the four β-propeller domains (green) and four EGF-like repeats (cyan) indicated. Binding sites for specific Wnt ligands, various inhibitors, and antibodies generated to these molecules [72, 73] are also noted. Color schemes are consistent with those shown in Panels B and C. This figure is adapted from Cheng et al. [74] (B) A ribbon diagram of the LRP6 E1 structure in complex with the DKK1 peptide NSNAIKN (PDB code 3SOQ) is shown. The peptide sequence is colored in magenta with the NAIK motif shown as a stick model. (C) The Lrp6 E1E2 structure in complex with the DKK1_C domain is represented by a ribbon diagram (PDB code 3S94). (D) A ribbon diagram depicts the Lrp6 E3E4 structure in complex DKK1_C domain (PDB code 3S2K). The DKK1_C structure is colored in magenta with disulfide bonds shown in yellow. Only one copy of DKK1_C and Lrp6E3E4 corresponding to the interface A is shown.