Identification of PLOD2 as Telopeptide Lysyl Hydroxylase, an Important Enzyme in Fibrosis

Abstract

The hallmark of fibrotic processes is an excessive accumulation of collagen. The deposited collagen shows an increase in pyridinoline cross-links, which are derived from hydroxylated lysine residues within the telopeptides. This change in cross-linking is related to irreversible accumulation of collagen in fibrotic tissues. The increase in pyridinoline cross-links is likely to be the result of increased activity of the enzyme responsible for the hydroxylation of the telopeptides (telopeptide lysyl hydroxylase, or TLH). Although the existence of TLH has been postulated, the gene encoding TLH has not been identified. By analyzing the genetic defect of Bruck syndrome, which is characterized by a pyridinoline deficiency in bone collagen, we found two missense mutations in exon 17 of PLOD2, thereby identifying PLOD2 as a putative TLH gene. Subsequently, we investigated fibroblasts derived from fibrotic skin of systemic sclerosis (SSc) patients and found that PLOD2 mRNA is highly increased indeed. Furthermore, increased pyridinoline cross-link levels were found in the matrix deposited by SSc fibroblasts, demonstrating a clear link between mRNA levels of the putative TLH gene (PLOD2) and the hydroxylation of lysine residues within the telopeptides. These data underscore the significance of PLOD2 in fibrotic processes.The biosynthesis of collagen molecules involves several intracellular post-translational modifications followed by excretion and extracellular aggregation of the collagen molecules into fibrils, which are subsequently stabilized by intermolecular cross-links (1, 2). Two related routes are responsible for the formation of these collagen cross-links, namely the allysine route, in which a lysine residue in the telopeptide is converted by lysyl oxidase into the aldehyde allysine, and the hydroxyallysine route, in which a hydroxylysine residue in the telopeptide is converted into the aldehyde hydroxyallysine. Subsequently, the allysine or the hydroxyallysine reacts with a Lys or Hyl residue in the triple helix to form di-, tri-, or tetrafunctional cross-links (3-6). The mature cross-links hydroxylysylpyridinoline or lysylpyridinoline are formed via the hydroxyallysine route and occur in a variety of connective tissues such as bone, tendon, ligaments, and cartilage (7). In contrast, collagen in the skin is mainly cross-linked via the allysine route. Interestingly, in fibrotic skin (lipodermatosclerosis, keloid) and organ fibrosis (lung, liver), which is characterized by an excessive accumulation of collagen, an increase in crosslinks derived from the hydroxyallysine route is found (8 -15). It has been shown that the amount of hydroxyallysine-derived cross-links is related to the irreversible accumulation of collagen in fibrotic tissues, indicating that collagen containing hydroxyallysine-derived cross-links is more difficult to degrade than collagen containing allysine-derived cross-links (10 -12, 14, 15). Inhibition of the formation of hydroxyallysine-derived cross-links in fibrosis is therefore likely to resu...

The IκB kinase inhibitor ACHP strongly attenuates TGFβ1‐induced myofibroblast formation and collagen synthesis

“…The substrate specificity of LH2a is unknown, but most likely not the telopeptides. A higher expression of LH2 has been reported in fibrosis (specifically: LH2b) 32, 33, 34, 36, explaining the higher level of pyridinoline cross‐links. How the expression of PLOD1 , PLOD2 and PLOD3 (encoding LH1, LH2 and LH3 respetively) is regulated is essentially unknown.…”

Osteogenesis imperfecta

“…Collagen secreted from FKBP65 defective cells shows normal 3 hydroxylation of proline 986 of the α1(I) chain and normal to only slightly overmodified helical modifications, but C telopeptide lysine residues that are crucial for collagen crosslinking 18,79 are severely under hydroxylated 76 . The hydroxylation of C telopeptide lysine residues is catalysed by LH2, deficiency of which mainly leads to Bruck syndrome, but also to forms of recessive osteogenesis imperfecta without contrac tures [80][81][82] . How FKBP65 decreases LH2 activity is not yet understood.…”

Premature aggregation of type IV collagen and early lethality in lysyl hydroxylase 3 null mice

“…Many patients with the kyphoscoliotic type of Ehlers-Danlos syndrome with mutations in the LH1 gene have a virtual absence of hydroxylysine in the collagens of their skin and some other tissues and yet have a relatively mild disease phenotype (7)(8)(9). The clinical manifestations in the kyphoscoliotic type of Ehlers-Danlos syndrome and in Bruck syndrome, where mutations in the LH2 gene were recently identified (10), are believed to be due to impaired stability of crosslinks in fibrillar collagens. Our analyses show that functions of the third isoform, LH3, are crucial with regard to both embryonic development and type IV collagen assembly.…”

“…Mutations in the LH1 gene lead to the kyphoscoliotic type of Ehlers-Danlos syndrome, which is characterized by scoliosis, joint laxity, skin fragility, ocular manifestations, and severe muscle hypotonia (7)(8)(9). Mutations in the LH2 gene have recently been reported in two families with Bruck syndrome, which is characterized by fragile bones, joint contractures, scoliosis, and osteoporosis (10).…”