The Pro-regions of Lysyl Oxidase and Lysyl Oxidase-like 1 Are Required for Deposition onto Elastic Fibers

Abstract

These studies were undertaken to determine how lysyl oxidase (LOX) and lysyl oxidase like-1 (LOXL) enzymes are targeted to their substrates in the extracellular matrix. Full-length LOX/LOXL and constructs containing just the pro-regions of each enzyme localized to elastic fibers when expressed in cultured cells. However, the LOXL catalytic domain without the pro-region was secreted into the medium but did not associate with matrix. Ligand blot and mammalian two-hybrid assays confirmed an interaction between tropoelastin and the pro-regions of both LOX and LOXL. Immunofluorescence studies localized both enzymes to elastin at the earliest stages of elastic fiber assembly. Our results showed that the proregions of LOX and LOXL play a significant role in directing the deposition of both enzymes onto elastic fibers by mediating interactions with tropoelastin. These findings confirmed that an important element of substrate recognition lies in the pro-domain region of the molecule and that the pro-form of the enzyme is what initially interacts with the matrix substrate. These results have raised the interesting possibility that sequence differences between the prodomain of LOX and LOXL account for some of the functional differences observed for the two enzymes.Production of a mature and functional elastic fiber is a complex process that is only partially understood. Monomers of elastin (tropoelastin) are cross-linked in the extracellular space by one or more members of the lysyl oxidase (LO) 3 gene family to form an elastin polymer, which is the functional form of the mature protein. Fibrillin-containing microfibrils are thought to play an important role in the assembly process by serving as a scaffold for aligning cross-linking domains within tropoelastin. Recently, several other proteins, such as members of the fibulin and emilin families, have been suggested to play a role in elastic fiber formation, although their exact function has not yet been determined (1).LOs are extracellular copper-requiring enzymes that catalyze the cross-linking of collagen and elastin through oxidative deamination of lysine or hydroxylysine side chains. The resultant allysine residues can then spontaneously condense with vicinal peptidyl aldehydes or with ⑀-amino groups of peptidyl lysines to generate covalent cross-linkages.There are five members of the LO family: lysyl oxidase (LOX) and lysyl oxidase-like 1-4 (LOXL 1-4) (reviewed in Ref.2). The C-terminal region of all of the LO family members contains the elements required for catalytic activity (the copper binding site, tyrosyl and lysyl residues that contribute to the carbonyl cofactor, and 10 cysteine residues), and the high sequence homology in this region suggests that all family members share a common enzymatic mechanism. The N-terminal regions, in contrast, show the greatest variability in size and sequence.The genes for LOX and LOXL have a similar exon structure consisting of seven exons, five of which (exons 2-6) are of similar size and encode proteins with 76% amino acid iden...

New insights into elastic fiber assembly

“…LOXL1 specifically localizes to sites of elastogenesis and interacts with fibulin-5 (Liu et al, 2004). The pro region plays a role in directing LOX and LOXL1 to elastic fibers in vitro, and sequence differences in the pro regions may account for some of the functional differences between the enzymes (Thomassin et al, 2005).…”

The Lysyl Oxidase Pro-peptide Attenuates Fibronectin-mediated Activation of Focal Adhesion Kinase and p130Cas in Breast Cancer Cells

“…By this mechanism it would enhance the enzymatic activity of LOX and might counteract the tumor suppressor function of the LOX-PP. Moreover, it has been demonstrated that LOX-PP plays a crucial role in the deposition of the enzyme onto elastic fibers by interacting with tropoelastin (39). During tumor progression, the microenvironment undergoes significant changes.…”

Microarray gene expression analysis in atrophying rainbow trout muscle: a unique nonmammalian muscle degradation model

“…The posttranslational list also includes protein disulfide-isomerase A3 and protein-lysine 6-oxidase. The latter is an extracellular enzyme that catalyzes the cross-linking of collagen and elastin through oxidative deamination of lysine or hydroxylysine side chains (68). Weakening of the cross-linking of these proteins may further explain the decreased muscle integrity and quality of RBT because of the vitellogenesis effect.…”