Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma

Chen, Yulong; Terajima, Masahiko; Yang, Yanan; Sun, Li; Ahn, Young Ho; Paňková, Daniela; Puperi, Daniel S.; Watanabe, Takeshi; Kim, Min P.; Blackmon, Shanda H.; Rodríguez, Jaime; Liu, Hui; Behrens, Carmen; Wistuba, Ignacio I.; Minelli, Rosalba; Scott, Kenneth L.; Sanchez-Adams, Johannah; Guilak, Farshid; Pati, Debananda; Thilaganathan, Nishan; Burns, Alan R.; Creighton, Chad J.; Martínez, Elisabeth D.; Żal, Tomasz; Grande-Allen, K. Jane; Yamauchi, Mitsuo; Kurie, Jonathan M.

doi:10.1172/jci74725

Cited by 146 publications

(197 citation statements)

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“…Considering that only dimers seem to exhibit LH activity, our data provide a molecular explanation as to how mutations in FKBP10 result in an overall dramatic decrease in lysyl hydroxylation levels of collagen telopeptides, leading to diseases such as Bruck syndrome, type XI osteogenesis imperfecta, and Kuskokwim syndrome. It should be noted that a high level of pyridinoline crosslinks has been reported to be causally involved in fibrosis and tumor metastasis (28,(50)(51)(52)(53)(54). The increased stiffening of the matrix from accumulating cross-linked collagen is directly linked to the progression of both types of pathologies.…”

Section: Discussionmentioning

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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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%

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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“…The dimerization motif (amino acids 562ϳ568) on LH2 was deleted by overlapping PCR to generate the LH2-DD-G1 and LH2-DD-G2 expression plasmids. The pLKO.1_scramble and pLKO.1_LH2-SH5 plasmids were constructed as described previously (21). To create the pEF-bsr plasmid for the rescue experiment, we modified the pLVX-Puro plasmid by replacing the CMV promoter with the EF1␣ promoter, replacing the puromycin-resistant gene with the blasticidinresistant gene, and adding the SbfI and NotI cutting sites after the XbaI site.…”

Section: Methodsmentioning

confidence: 99%

“…In lung cancer, high LH2 expression increases the amount of HLCCs in tumor stroma (21). Moreover, LH2 has been detected in the secretome of breast cancer cells (23), which raises the possibility that LH2 has an extracellular function.…”

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confidence: 99%

Lysyl Hydroxylase 2 Is Secreted by Tumor Cells and Can Modify Collagen in the Extracellular Space

Chen

Guo

Terajima

et al. 2016

Journal of Biological Chemistry

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Edited by Amanda FosangLysyl hydroxylase 2 (LH2) catalyzes the hydroxylation of lysine residues in the telopeptides of fibrillar collagens, which leads to the formation of stable collagen cross-links. Recently we reported that LH2 enhances the metastatic propensity of lung cancer by increasing the amount of stable hydroxylysine aldehyde-derived collagen cross-links (HLCCs), which generate a stiffer tumor stroma (Chen, Y., et al. (2015) J. Clin. Invest. 125, 125, 1147-1162). It is generally accepted that LH2 modifies procollagen ␣ chains on the endoplasmic reticulum before the formation of triple helical procollagen molecules. Herein, we report that LH2 is also secreted and modifies collagen in the extracellular space. Analyses of lung cancer cell lines demonstrated that LH2 is present in the cell lysates and the conditioned media in a dimeric, active form in both compartments. LH2 colocalized with collagen fibrils in the extracellular space in human lung cancer specimens and in orthotopic lung tumors generated by injection of a LH2-expressing human lung cancer cell line into nude mice. LH2 depletion in MC3T3 osteoblastic cells impaired the formation of HLCCs, resulting in an increase in the unmodified lysine aldehyde-derived collagen cross-link (LCC), and the addition of recombinant LH2 to the media of LH2-deficient MC3T3 cells was sufficient to rescue HLCC formation in the extracellular matrix. The finding that LH2 modifies collagen in the extracellular space challenges the current view that LH2 functions solely on the endoplasmic reticulum and could also have important implications for cancer biology.Collagens are the main component of the extracellular matrix and the most abundant protein in mammals (1). Of the 29 types of collagen identified, fibrillar type I collagen is the most abundant and important in providing tissues and organs with form and mechanical strength. Type I collagen biosynthesis involves a series of post-translational modifications of nascent procollagen ␣ chains on the endoplasmic reticulum; for example, specific lysine (Lys) and proline (Pro) residues undergo hydroxylation, and hydroxylysine (Hyl) residues undergo mono-and di-glycosylation. In the extracellular space, telopeptidyl Lys and Hyl residues on collagen molecules are oxidatively deaminated by lysyl oxidases (LOXs), 4 producing the reactive aldehydes Lys ald and Hyl ald , respectively, which initiates a series of condensation reactions to form various covalent intermolecular cross-links involving juxtaposed Lys, Hyl, and histidine (His) residues on the neighboring molecules, resulting in the formation of Hyl ald -derived collagen cross-links (HLCCs) (2).HLCCs are generated through a series of intermediate steps involving multiple enzymatic reactions. For example, HLCCs are produced through the divalent iminium cross-links dehydro-hydroxylysinonorleucine (deH-HLNL) when paired with a juxtaposed Lys residue (i.e. Hyl ald XLys) on a neighboring molecule, and deH-dihydroxylysinonorlecine (deH-DHLNL) with a Hyl residue (i.e. Hyl ald X...

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“…It is now clear that defective Lys modifications of collagen cause and/or are associated with a broad range of connective tissue disorders, including Ehlers-Danlos syndrome type VIA (3), bronchopulmonary dysplasia (4), Kuskokwim syndrome (5), Bruck syndrome (6,7), fibrosis (8), disuse osteoporosis (9), and cancer progression (10,11). Our recent finding showed that a switch of collagen cross-link phenotype may regulate tumor cell invasive and metastatic propensities (12).…”

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confidence: 99%

Cyclophilin-B Modulates Collagen Cross-linking by Differentially Affecting Lysine Hydroxylation in the Helical and Telopeptidyl Domains of Tendon Type I Collagen

Terajima

Taga

Chen

et al. 2016

Journal of Biological Chemistry

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Covalent intermolecular cross-linking provides collagen fibrils with stability. The cross-linking chemistry is tissue-specific and determined primarily by the state of lysine hydroxylation at specific sites. A recent study on cyclophilin B (CypB) null mice, a model of recessive osteogenesis imperfecta, demonstrated that lysine hydroxylation at the helical cross-linking site of bone type I collagen was diminished in these animals (Cabral, W. A., Perdivara, I., Weis, M., Terajima, M., Blissett, A. R., Chang, W., Perosky, J. E., Makareeva, E. N., Mertz, E. L., Leikin, S., Tomer, K. B., Kozloff, K. M., Eyre, D. R., Yamauchi, M., and Marini, J. C. (2014) PLoS Genet. 10, e1004465). However, the extent of decrease appears to be tissue-and molecular site-specific, the mechanism of which is unknown. Here we report that although CypB deficiency resulted in lower lysine hydroxylation in the helical cross-linking sites, it was increased in the telopeptide cross-linking sites in tendon type I collagen. This resulted in a decrease in the lysine aldehyde-derived cross-links but generation of hydroxylysine aldehyde-derived cross-links. The latter were absent from the wild type and heterozygous mice. Glycosylation of hydroxylysine residues was moderately increased in the CypB null tendon. We found that CypB interacted with all lysyl hydroxylase isoforms (isoforms 1-3) and a putative lysyl hydroxylase-2 chaperone, 65-kDa FK506-binding protein. Tendon collagen in CypB null mice showed severe size and organizational abnormalities. The data indicate that CypB modulates collagen cross-linking by differentially affecting lysine hydroxylation in a site-specific manner, possibly via its interaction with lysyl hydroxylases and associated molecules. This study underscores the critical importance of collagen post-translational modifications in connective tissue formation.Collagens comprise a large family of structurally related extracellular matrix proteins (1). Among all of the genetic types of collagen identified, fibrillar type I collagen is the most abundant, providing tissues and organs with form and stability. It is a heterotrimeric molecule composed of two ␣1 chains and one ␣2 chain forming a long uninterrupted triple helix with short non-helical domains (telopeptide) at both N and C termini. One of the functionally important characteristics of collagen is its unique, sequential post-translational modifications of Lys residues. The modifications include hydroxylation and monoand diglycosylation of hydroxylysine (Hyl), 3 and oxidative deamination of Lys and Hyl in the N-and C-telopeptides followed by extensive covalent intermolecular cross-linking (2). It is now clear that defective Lys modifications of collagen cause and/or are associated with a broad range of connective tissue disorders, including Ehlers-Danlos syndrome type VIA (3), bronchopulmonary dysplasia (4), Kuskokwim syndrome (5), Bruck syndrome (6, 7), fibrosis (8), disuse osteoporosis (9), and cancer progression (10, 11). Our recent finding showed that a switch of collage...

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Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma

Cited by 146 publications

References 67 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

Lysyl Hydroxylase 2 Is Secreted by Tumor Cells and Can Modify Collagen in the Extracellular Space

Cyclophilin-B Modulates Collagen Cross-linking by Differentially Affecting Lysine Hydroxylation in the Helical and Telopeptidyl Domains of Tendon Type I Collagen

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