Human lysyl oxidase-like 2

Abstract

Lysyl oxidase like-2 (LOXL2) belongs to the lysyl oxidase (LOX) family, which comprises Cu(2+)- and lysine tyrosylquinone (LTQ)-dependent amine oxidases. LOXL2 is proposed to function similarly to LOX in the extracellular matrix (ECM) by promoting crosslinking of collagen and elastin. LOXL2 has also been proposed to regulate extracellular and intracellular cell signaling pathways. Dysregulation of LOXL2 has been linked to many diseases, including cancer, pro-oncogenic angiogenesis, fibrosis and heart diseases. In this review, we will give an overview of the current understandings and hypotheses regarding the molecular functions of LOXL2.

Keywords: Cell signaling; Extracellular matrix; Lysine tyrosylquinone (LTQ); Lysyl oxidase family; Lysyl oxidase like-2; Tumor metastasis/invasion.

Figure 1:. Cartoon diagram of the LOX family of proteins.

Based on their N-termini, the LOX family of proteins is divided into two subgroups. LOX and LOXL1 constitute one subfamily possessing a highly basic propeptide sequence that is proteolytically removed by bone morphogenetic protein-1 (BMP-1). The exact site of BMP-1 cleavage in LOXL1 is not defined. Additionally, LOXL1 has a proline-rich domain within its propeptide. The other subfamily comprises LOXL2–4, each of which contains 4 scavenger receptor cysteine-rich (SRCR) domains instead of a propeptide. All LOX family members possess a C-terminal amine oxidase catalytic domain containing a putative copper-binding site (His-X-His-X-His), as well as an LTQ cofactor formed from conserved Lys and Tyr residues by post-translational modification. Each member also has an N-terminal secretion signal. LOX and LOXL2–4 contain predicted N-linked glycosylation sites (i.e. Asn-X-Ser/Thr). From (50).

Figure 2:. Structure of the LTQ cofactor and proposed pathway of LTQ cofactor biogenesis.

A) Structure of LTQ, the tyrosine-derived cofactor of LOXL2. B) The proposed mechanism for the biogenesis of the LTQ cofactor. Steps: (i) autocatalytic oxidation of the precursor peptidyl tyrosine residue by O2 and Cu²⁺; (ii) O2 oxidation of dihydroxyphenylalanine (DOPA) to dopaquinone (DPQ); (iii) 1,4-addition of the ε-amino group of the side chain of a peptidyl lysine residue (Lys653 in human LOXL2) to DPQ yields the reduced form of LTQ, where neutral amino side chain of Lys653 is a nucleophile; (iv) Oxidation of reduced form of LTQ to LTQ by O₂. Adapted from (9,41).

Figure 3.. X-ray crystal structure of LTQ-like quinone in D298K mutant of a bacterial CAO.

Active site structures of mature D298K-AGAO and the putative DPQ intermediate detected during X-ray snapshot analysis of TPQ biogenesis in WT-AGAO. (A) DPQ intermediate (PDB: 1IVV)(88), (B) D298K (PDB: 2YX9). Cu²⁺ is shown as an orange sphere, water molecules are shown as light-blue spheres, hydrogen bonding interactions are represented by blue lines, and ligand interactions are represented by purple lines. Val282 and Asn381 (white) form the edges of a wedge-shaped pocket. Hydrogen bonding distances are denoted in angstroms. From (42).

Figure 4:. Epithelial-to-Mesenchymal transition (EMT).

EMT is a process in which epithelial cells are transformed into mesenchymal cells. Changes in the tumor microenvironment (e.g. growth factors, cytokines, and extracellular matrix components) have been shown to induce EMT. During EMT, cells downregulate expression of cell-cell adhesion and cell polarity proteins, while upregulating expression of proteins that confer invasive and migratory traits. Adapted from (61).

Figure 5:. Proposed roles for LOXL2 in breast cancer metastasis and invasion.

A) Proposed mechanism whereby secreted LOXL2 induces ECM stiffening to activate oncogenic signaling pathways. B) Proposed mechanism whereby intracellular LOXL2 regulates Snail1 or H3K4me3 to repress E-cadherin expression, eventually leading to EMT.

Figure 6:. Proposed mechanism of LOXL2 oxidation and stabilization of Snail1.

Intracellular LOXL2 is proposed to deaminate Lys98 and Lys137 in the SNAG domain of Snail1. Following the modification by LOXL2, an undetermined conformational change in Snail1 inhibits phosphorylation by GSK3β and ultimately suppresses the degradation of Snail1.

Figure 7:. Proposed mechanisms for regulating methylated H3K4.

A) A novel mechanism proposed by Herranz et al. whereby LOXL2 deaminates H3K4(me3) via two steps: formation of an alcohol via deamination, and subsequent oxidation of alcohol to aldehyde. From (79). B) A scheme of the mechanism by which a Jumonji C (JMJC)-containing lysine demethylase, which is an Fe²⁺- and α-ketoglutarate-dependent dioxygenase, catalyzes demethylation of H3K4me3. Adapted from (84). C) A proposed mechanism for lysine side chain oxidation by LOXL2.

Scheme 1:. LOX-initiated crosslink formation in tropocollagen and tropoelastin.

A) Lysyl oxidase catalyzes the oxidative deamination of lysine and hydroxylysine residues in tropocollagen and tropoelastin. For simplicity, only lysine residues are shown in this scheme. B) The product allysine residues spontaneously react with other allysine or lysine residues via aldol condensation or Schiff base formation. C) The bifunctional condensation products can further crosslink to form tri-, tetra-, or even pentafunctional crosslinks. Depicted is desmosine, a common tetrafunctional crosslink found in elastin. Adapted from (87).

Scheme 2:. LTQ-LTI tautomerism.

The LTQ-like quinone (λ_max=504 nm) formed in D298KAGAO is in equilibrium with its iminoquinone tautomer (LTI, λ_max=454 nm). The LTI form is thermodynamically favored as it is stabilized by the hydrogen bonding interaction with the conserved Tyr284 in the active site. When the hydrogen bonding interaction was disrupted via site-directed mutagenesis (i.e. D298K/Y284F-AGAO), only the LTQ-like form was detected. From (42).