Glycosylation catalyzed by lysyl hydroxylase 3 is essential for basement membranes

Ruotsalainen, Heli; Sipilä, Laura; Vapola, Miia; Sormunen, Raija; Salo, Antti M.; Uitto, Lahja; Mercer, Derry K.; Robins, Simon P.; Risteli, Maija; Aszódi, Attila; Fässler, Reinhard; Myllylä, Raili

doi:10.1242/jcs.02780

Cited by 103 publications

(121 citation statements)

References 52 publications

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“…Mutations in LH2 (Bruck syndrome, type 2) results in severe underhydroxylation of the telopeptides of collagen type I in bone, whereas the lysyl hydroxylation level of the triple helix remains unaffected, indicating that LH2 is a telopeptide LH, and that LH1 and LH3 are triple-helical LHs. Indeed, mutations in LH1 (EhlersDanlos syndrome, type VIA) result in an underhydroxylation of the triple helix, but the telopeptides remain unaffected, and a knockout model of LH3 has shown unaffected telopeptides in bone (14,15). Therefore, it was surprising that mutations in FKBP10 (Bruck syndrome, type 1) resulted in a similar phenotype and in the same biochemical defect as seen for mutations in LH2.…”

Section: Discussionmentioning

confidence: 99%

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Disentangling mechanisms involved in collagen pyridinoline cross-linking: The immunophilin FKBP65 is critical for dimerization of lysyl hydroxylase 2

Gjaltema

Stoel

Boersema

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Collagens are subjected to extensive posttranslational modifications, such as lysine hydroxylation. Bruck syndrome (BS) is a connective tissue disorder characterized at the molecular level by a loss of telopeptide lysine hydroxylation, resulting in reduced collagen pyridinoline cross-linking. BS results from mutations in the genes coding for lysyl hydroxylase (LH) 2 or peptidyl-prolyl cis-trans isomerase (PPIase) FKBP65. Given that the immunophilin FKBP65 does not exhibit LH activity, it is likely that LH2 activity is somehow dependent on FKPB65. In this report, we provide insights regarding the interplay between LH2 and FKBP65. We found that FKBP65 forms complexes with LH2 splice variants LH2A and LH2B but not with LH1 and LH3. Ablating the catalytic activity of FKBP65 or LH2 did not affect complex formation. Both depletion of FKBP65 and inhibition of FKBP65 PPIase activity reduced the dimeric (active) form of LH2 but did not affect the binding of monomeric (inactive) LH2 to procollagen Iα1. Furthermore, we show that LH2A and LH2B cannot form heterodimers with each other but are able to form heterodimers with LH1 and LH3. Collectively, our results indicate that FKBP65 is linked to pyridinoline cross-linking by specifically mediating the dimerization of LH2. Moreover, FKBP65 does not interact with LH1 and LH3, explaining why in BS triple-helical hydroxylysines are not affected. Our results provide a mechanistic link between FKBP65 and the loss of pyridinolines and may hold the key to future treatments for diseases related to collagen cross-linking anomalies, such as fibrosis and cancer.collagen cross-linking | lysyl hydroxylase | FKBP65 | Bruck syndrome | fibrosis C ollagen I is an essential component of the extracellular matrix (ECM) of tissues such as bone and skin, and is involved in a wide variety of biological processes. A deregulated synthesis of collagen type I results in pathologies ranging from severe bone and skin anomalies to fibrosis (1-5). Hydroxylation of specific lysine (Lys) residues into 5-hydroxylysine (Hyl) is performed by lysyl hydroxylases (LHs), also known as the procollagen-lysine, 2-oxoglutarate 5-dioxygenase (PLOD) family. Collagens deposited in the ECM are stabilized by the formation of intermolecular crosslinks by members of the lysyl oxidase family, LOX and LOXL (6). Two collagen cross-linking pathways have been identified, the allysine route and the hydroxyallysine route (7). In the allysine route, a telopeptide Lys is oxidized by lysyl oxidases into an aldehyde; in the hydroxyallysine route, this occurs with a telopeptide Hyl. In turn, the reactive aldehydes interact with the Lys or Hyl residues in the helical part of collagen to form difunctional and finally trifunctional cross-links (8-10). The trifunctional cross-links derived from the hydroxyallysine route are referred to as pyridinolines. Collagens cross-linked by means of pyridinolines are difficult to degrade (11-13).The LH family consists of three individual members, designated LH1, LH2, and LH3. Hydroxylation of Lys present...

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Section: Discussionmentioning

confidence: 99%

“…Hydroxylation of Lys present in the Gly-X-Lys sequence in the helical domain of collagen is carried out by LH1 and LH3 (14,15), whereas LH2 hydroxylates Lys residues in the telopeptides of collagen, where such a sequence motif is not present (16)(17)(18). LH2 consists of two splice variants, LH2A and LH2B (19), with LH2B containing an extra exon known as 13A.…”

mentioning

confidence: 99%

Disentangling mechanisms involved in collagen pyridinoline cross-linking: The immunophilin FKBP65 is critical for dimerization of lysyl hydroxylase 2

Gjaltema

Stoel

Boersema

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Defects of triple helix formation because of altered posttranslational modifications of collagen often impair the trafficking of collagens (7,30). Under physiological conditions, only triple helical collagen is secreted, indicating that collagen secretion is directly proportional to triple helix formation (31).…”

Section: Glt25d1 and Glt25d2 Inactivation In Osteosarcomamentioning

confidence: 99%

Collagen Accumulation in Osteosarcoma Cells lacking GLT25D1 Collagen Galactosyltransferase

Baumann

Hennet

2016

Journal of Biological Chemistry

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Collagen is post-translationally modified by prolyl and lysyl hydroxylation and subsequently by glycosylation of hydroxylysine. Despite the widespread occurrence of the glycan structure Glc(␣1-2)Gal linked to hydroxylysine in animals, the functional significance of collagen glycosylation remains elusive. To address the role of glycosylation in collagen expression, folding, and secretion, we used the CRISPR/Cas9 system to inactivate the collagen galactosyltransferase GLT25D1 and GLT25D2 genes in osteosarcoma cells. Loss of GLT25D1 led to increased expression and intracellular accumulation of collagen type I, whereas loss of GLT25D2 had no effect on collagen secretion. Inactivation of the GLT25D1 gene resulted in a compensatory induction of GLT25D2 expression. Loss of GLT25D1 decreased collagen glycosylation by up to 60% but did not alter collagen folding and thermal stability. Whereas cells harboring individually inactivated GLT25D1 and GLT25D2 genes could be recovered and maintained in culture, cell clones with simultaneously inactive GLT25D1 and GLT25D2 genes could be not grown and studied, suggesting that a complete loss of collagen glycosylation impairs osteosarcoma cell proliferation and viability.

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“…Although their biological function still remains somewhat unclear (73), they are believed to play a role in modulating the structural stability of collagen (74,75). Indeed, collagen glycosylation plays a role in bone mineralization; however, overglycosylation has been shown to be associated with disorders such as osteogenesis imperfecta (76), chondrodysplasias (77), rheumatoid arthritis (78), postmenopausal osteoporosis, osteosarcoma, osteofibrous dysplasia, and Kashin-Beck disease (73)-further suggesting a role for these particular modifications in modulating collagen architecture.…”

Section: Table I Proteins Identified In Ice Age Bison Skull (Identifimentioning

confidence: 99%

Preserved Proteins from Extinct Bison latifrons Identified by Tandem Mass Spectrometry; Hydroxylysine Glycosides are a Common Feature of Ancient Collagen

Hill

Wither

Nemkov

et al. 2015

Molecular & Cellular Proteomics

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Bone samples from several vertebrates were collected from the Ziegler Reservoir fossil site, in Snowmass Village, Colorado, and processed for proteomics analysis. The specimens come from Pleistocene megafauna Bison latifrons, dating back ϳ120,000 years. Proteomics analysis using a simplified sample preparation procedure and tandem mass spectrometry (MS/MS) was applied to obtain protein identifications. Several bioinformatics resources were used to obtain peptide identifications based on sequence homology to extant species with annotated genomes. With the exception of soil sample controls, all samples resulted in confident peptide identifications that mapped to type I collagen. In addition, we analyzed a specimen from the extinct B. latifrons that yielded peptide identifications mapping to over 33 bovine proteins. Our analysis resulted in extensive fibrillar collagen sequence coverage, including the identification of posttranslational modifications. Hydroxylysine glucosylgalactosylation, a modification thought to be involved in collagen fiber formation and bone mineralization, was identified for the first time in an ancient protein dataset. Meta-analysis of data from other studies indicates that this modification may be common in well-preserved prehistoric samples. Additional peptide sequences from extracellular matrix (ECM) and non-ECM proteins have also been identified for the first time in ancient tissue samples. These data provide a framework for analyzing ancient protein signatures in wellpreserved fossil specimens, while also contributing novel insights into the molecular basis of organic matter preservation. As such, this analysis has unearthed common posttranslational modifications of collagen that may assist in its preservation over time. During the last decade, paleontology and taphonomy (the study of decaying organisms over time and the fossilization processes) have begun to overlap with the field of proteomics to shed new light on preserved organic matter in fossilized bones (1-4). These bones represent a time capsule of ancient biomolecules, owing to their natural resistance to post mortem decay arising from a unique combination of mechanical, structural, and chemical properties (4 -7).Although bones can be cursorily described as a composite of collagen (protein) and hydroxyapatite (mineral), fossilized bones undergo three distinct diagenesis pathways: (i) chemical deterioration of the organic phase; (ii) chemical deterioration of the mineral phase; and (iii) (micro)biological attack of the composite (6). In addition, the rate of these degradation pathways are affected by temperature, as higher burial temperatures have been shown to accelerate these processes (6,8). Though relatively unusual, the first of these three pathways results in a slower deterioration process, which is more generally mitigated under (6) specific environmental constraints, such as geochemical stability (stable temperature and acidity) that promote bone mineral preservation. Importantly, slower deterioration results in more pre...

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Glycosylation catalyzed by lysyl hydroxylase 3 is essential for basement membranes

Cited by 103 publications

References 52 publications

Disentangling mechanisms involved in collagen pyridinoline cross-linking: The immunophilin FKBP65 is critical for dimerization of lysyl hydroxylase 2

Disentangling mechanisms involved in collagen pyridinoline cross-linking: The immunophilin FKBP65 is critical for dimerization of lysyl hydroxylase 2

Collagen Accumulation in Osteosarcoma Cells lacking GLT25D1 Collagen Galactosyltransferase

Preserved Proteins from Extinct Bison latifrons Identified by Tandem Mass Spectrometry; Hydroxylysine Glycosides are a Common Feature of Ancient Collagen

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