FKBP65-dependent peptidyl-prolyl isomerase activity potentiates the lysyl hydroxylase 2-driven collagen cross-link switch

Bruck Syndrome is a connective tissue disease associated with inactivating mutations in lysyl hydroxylase 2 (LH2/PLOD2) or FK506 binding protein 65 (FKBP65/FKBP10). However, the functional relationship between LH2 and FKBP65 remains unclear. Here, we postulated that peptidyl prolyl isomerase (PPIase) activity of FKBP65 positively modulates LH2 enzymatic activity and is critical for the formation of hydroxylysine-aldehyde derived intermolecular collagen cross-links (HLCCs). To test this hypothesis, we analyzed collagen cross-links in Fkbp10-null and –wild-type murine embryonic fibroblasts. Although LH2 protein levels did not change, FKBP65 deficiency significantly diminished HLCCs and increased the non-hydroxylated lysine-aldehyde–derived collagen cross-links (LCCs), a pattern consistent with loss of LH2 enzymatic activity. The HLCC-to-LCC ratio was rescued in FKBP65-deficient murine embryonic fibroblasts by reconstitution with wild-type but not mutant FKBP65 that lacks intact PPIase domains. Findings from co-immunoprecipitation, protein-fragment complementation, and co-immunofluorescence assays showed that LH2 and FKBP65 are part of a common protein complex. We conclude that FKBP65 regulates LH2-mediated collagen cross-linking. Because LH2 promotes fibrosis and cancer metastasis, our findings suggest that pharmacologic strategies to target FKBP65 and LH2 may have complementary therapeutic activities

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

Lysyl 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 α 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 co-localized 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

Metal Bond Strength Regulation Enables Large-scale Synthesis of Intermetallic Nanocrystals for Practical Fuel Cells

Structurally ordered L10-PtM (M = Fe, Co, Ni, etc) intermetallic nanocrystals (iNCs), benefiting from the chemically ordered structure and higher stability, are one of the best electrocatalysts used for PEMFC. However, their practical development is greatly plagued by the challenge that high-temperature annealing (> 700 °C) has to be used for realizing disorder-order phase transition (DOPT) due to the high activation barrier (Ea), which always leads to severe particle sintering, morphology change, and makes it highly challenging for gram-scale preparation of desirable PtM iNCs. Here, we report a general low-melting-point metal induced bond strength weakening strategy to promote DOPT of PtM (M = Ni, Fe, Cu, Zn) alloy catalysts. We demonstrate that the introduction of Sn can reduce DOPT temperature to a record-low temperature (≤ 450 °C), which enables ten-gram-scale preparation of high-performance L10-PtM iNCs. X-ray spectroscopic studies, in-situ electron microscopy and theoretical calculations reveal that the Sn-facilitated DOPT mechanism at record-low temperature involves the weakened bond strength and reduced Ea via Sn doping, the formation and fast diffusion of low coordinated surface free atom, and subsequent L10 nucleation. Most importantly, the 15% Sn-doped L10-PtNi iNCs display outstanding performance in H2-air fuel cells with a high peak power density of 1.45 W cm-2 for Pt alloy catalysts and less than 25% activity loss after 30000 cycles at a quite low cathode Pt loading amount of 0.12 mg¬Pt cm-2, representing as one of the most efficient cathodic electrocatalyst for PEMFCs