Crystal structure of LRG1 and the functional significance of LRG1 glycan for LPHN2 activation
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The serum glycoprotein leucine-rich ɑ-2-glycoprotein 1 (LRG1), primarily produced by hepatocytes and neutrophils, is a multifunctional protein that modulates various signaling cascades, mainly TGFβ signaling. Serum LRG1 and neutrophil-derived LRG1 have different molecular weights due to differences in glycosylation, but the impact of the differential glycan composition in LRG1 on its cellular function is largely unknown. We previously reported that LRG1 can promote both angiogenic and neurotrophic processes under hyperglycemic conditions by interacting with LPHN2. Here, we determined the crystal structure of LRG1, identifying the horseshoe-like solenoid structure of LRG1 and its four N-glycosylation sites. In addition, our biochemical and cell-biological analyses found that the deglycosylation of LRG1, particularly the removal of glycans on N325, is critical for the high-affinity binding of LRG1 to LPHN2 and thus promotes LRG1/LPHN2-mediated angiogenic and neurotrophic processes in mouse tissue explants, even under normal glucose conditions. Moreover, the intracavernous administration of deglycosylated LRG1 in a diabetic mouse model ameliorated vascular and neurological abnormalities and restored erectile function. Collectively, these data indicate a novel role of LRG1 glycans as molecular switches that can tune the range of LRG1’s cellular functions, particularly the LRG1/LPHN2 signaling axis.
Abstract
The serum glycoprotein leucine-rich ɑ-2-glycoprotein 1 (LRG1), primarily produced by hepatocytes and neutrophils, is a multifunctional protein that modulates various signaling cascades, mainly TGFβ signaling. Serum LRG1 and neutrophil-derived LRG1 have different molecular weights due to differences in glycosylation, but the impact of the differential glycan composition in LRG1 on its cellular function is largely unknown. We previously reported that LRG1 can promote both angiogenic and neurotrophic processes under hyperglycemic conditions by interacting with LPHN2. Here, we determined the crystal structure of LRG1, identifying the horseshoe-like solenoid structure of LRG1 and its four N-glycosylation sites. In addition, our biochemical and cell-biological analyses found that the deglycosylation of LRG1, particularly the removal of glycans on N325, is critical for the high-affinity binding of LRG1 to LPHN2 and thus promotes LRG1/LPHN2-mediated angiogenic and neurotrophic processes in mouse tissue explants, even under normal glucose conditions. Moreover, the intracavernous administration of deglycosylated LRG1 in a diabetic mouse model ameliorated vascular and neurological abnormalities and restored erectile function. Collectively, these data indicate a novel role of LRG1 glycans as molecular switches that can tune the range of LRG1’s cellular functions, particularly the LRG1/LPHN2 signaling axis.
참고문헌 (54)
- Haupt, H., "Isolation and characterization of an unknown, leucinerich 3.1-S-alpha2-glycoprotein from human serum (author’s transl)", Hoppe Seylers Z. Physiol. Chem, 358, pp.639~646 (1977)
- Takahashi, N., "Periodicity of leucine and tandem repetition of a 24-amino acid segment in the primary structure of leucine-rich alpha 2-glycoprotein of human serum", Proc. Natl Acad. Sci. USA, 82, pp.1906~1910 (1985)
- Camilli, C., "LRG1 : an emerging player in disease pathogenesis", J. Biomed. Sci, 29, pp.6~ (2022)
- Wang, X., "LRG1 promotes angiogenesis by modulating endothelial TGF-beta signalling", Nature, 499, pp.306~311 (2013)
- Zou, Y., "Research Progress on Leucine-Rich Alpha-2 Glycoprotein 1 : A Review", Front. Pharmacol, 12, pp.809225~ (2021)
- Hong, Q., "LRG1 Promotes Diabetic Kidney Disease Progression by Enhancing TGF-beta-Induced Angiogenesis", J. Am. Soc. Nephrol, 30, pp.546~562 (2019)
- Haku, S., "Early Enhanced Leucine-Rich alpha-2-Glycoprotein-1 Expression in Glomerular Endothelial Cells of Type 2 Diabetic Nephropathy Model Mice", Biomed. Res. Int, 2018, pp.2817045~ (2018)
- Hisata, S., "Reversal of emphysema by restoration of pulmonary endothelial cells", J. Exp. Med, 218, pp.e20200938~ (2021)
- Singhal, M., "Temporal multi-omics identifies LRG1 as a vascular niche instructor of metastasis", Sci. Transl. Med, 13, pp.eabe6805~ (2021)
- Druhan, L. J., "Leucine Rich alpha-2 Glycoprotein : A Novel Neutrophil Granule Protein and Modulator of Myelopoiesis", PLoS One, 12, pp.e0170261~ (2017)
- Jemmerson, R., "Paradoxical Roles of Leucine-Rich alpha2-Glycoprotein-1 in Cell Death and Survival Modulated by Transforming Growth Factor-Beta 1 and Cytochrome c", Front. Cell. Dev. Biol, 9, pp.744908~ (2021)
- Codina, R., "Cytochrome c-induced lymphocyte death from the outside in : inhibition by serum leucine-rich alpha-2-glycoprotein-1", Apoptosis, 15, pp.139~152 (2010)
- Kajimoto, E., "Evaluation of leucine-rich alpha-2 glycoprotein as a biomarker of fetal infection", PLoS One, 15, pp.e0242076~ (2020)
- Shirai, R., "Up-regulation of the expression of leucine-rich alpha(2)-glycoprotein in hepatocytes by the mediators of acute-phase response", Biochem. Biophys. Res. Commun, 382, pp.776~779 (2009)
- Weivoda, S., "ELISA for human serum leucine-rich alpha-2-glycoprotein-1employing cytochrome c as the capturing ligand", J. Immunol. Methods, 336, pp.22~29 (2008)
- Chen, B., "Single Cell Transcriptome Data Analysis Defines the Heterogeneity of Peripheral Nerve Cells in Homeostasis and Regeneration", Front. Cell. Neurosci, 15, pp.624826~ (2021)
- Gao, Y., "LRG1 Promotes Keratinocyte Migration and Wound Repair through Regulation of HIF-1alpha Stability", J. Invest. Dermatol, 140, pp.455~464 (2020)
- Li, W., "Leucine-rich alpha-2-glycoprotein-1 promotes diabetic corneal epithelial wound healing and nerve regeneration via regulation of matrix metalloproteinases", Exp. Eye. Res, 196, pp.108060~ (2020)
- Liu, C., "A Multifunctional Role of Leucine-Rich-Alpha 2 Glycoprotein 1 in Cutaneous Wound Healing Under Normal and Diabetic Conditions", Diabetes, 69, pp.2467~2480 (2020)
- Yin Guo Nan, "Latrophilin-2 is a novel receptor of LRG1 that rescues vascular and neurological abnormalities and restores diabetic erectile function", Experimental and Molecular Medicine, 54, pp.1~13 (2022), 생화학분자생물학회[DOI]
- 박범석, "Human leucine-rich-alpha-2-glycoprotein1: purification, crystallization, and X-ray crystallographic analysis", Biodesign, 8(3), pp.60~63 (2020), 한국구조생물학회[DOI]
- McCoy, A. J., "Phaser crystallographic software", J. Appl. Crystallogr, 40, pp.658~674 (2007)
- Emsley, P., "Coot : model-building tools for molecular graphics", Acta Crystallogr. D, 60, pp.2126~2132 (2004)
- Adams, P. D., "PHENIX : building new software for automated crystallographic structure determination", Acta Crystallogr. D, 58, pp.1948~1954 (2002)
- Schrodinger, L. L. C., "The PyMOL Molecular Graphics System, v.1", pp.8~ (2015)
- Perner, C., "Protocol for dissection and culture of murine dorsal root ganglia neurons to study neuropeptide release", Star. Protoc, 2, pp.100333~ (2021)
- Jo, G., "Structural insights into the clustering and activation of Tie2 receptor mediated by Tie2 agonistic antibody", Nat. Commun, 12, pp.6287~ (2021)
- Lee, D. H., "Multi-paratopic VEGF decoy receptor have superior anti-tumor effects through anti-EGFRs and targeted anti-angiogenic activities", Biomaterials, 171, pp.34~45 (2018)
- Jin, H. R., "Functional and morphologic characterizations of the diabetic mouse corpus cavernosum : comparison of a multiple low-dose and a single highdose streptozotocin protocols", J. Sex. Med, 6, pp.3289~3304 (2009)
- Bella, J., "The leucine-rich repeat structure", Cell. Mol. Life. Sci, 65, pp.2307~2333 (2008)
- Ng, A. C., "Human leucine-rich repeat proteins : a genome-wide bioinformatic categorization and functional analysis in innate immunity", Proc. Natl Acad. Sci. USA, 108, pp.4631~4638 (2011)
- Seiradake, E., "Structural basis for cell surface patterning through NetrinGNGL interactions", Embo. J, 30, pp.4479~4488 (2011)
- Cummings, C., "Serum leucine-rich alpha-2-glycoprotein-1 binds cytochrome c and inhibits antibody detection of this apoptotic marker in enzyme-linked immunosorbent assay", Apoptosis, 11, pp.1121~1129 (2006)
- Shirai, R., "Autologous extracellular cytochrome c is an endogenous ligand for leucine-rich alpha2-glycoprotein and beta-type phospholipase A2 inhibitor", J. Biol. Chem, 285, pp.21607~21614 (2010)
- Kobe, B., "The leucine-rich repeat as a protein recognition motif", Curr. Opin. Struct. Biol, 11, pp.725~732 (2001)
- Kim, H. M., "Structural diversity of the hagfish variable lymphocyte receptors", J. Biol. Chem, 282, pp.6726~6732 (2007)
- Kim, H., "Structural basis for assembly and disassembly of the IGF", IGFBP, Nat. Commun, pp.2022, 13[DOI]
- Um, J. W., "Structural basis for LAR-RPTP", Slitrk complex-mediated synaptic adhesion, 2014, pp.5, 5423~
- Evans, R., "Protein complex prediction with AlphaFold-Multimer", biorxiv, ~ (2022)
- Kozakov, D., "The ClusPro web server for protein-protein docking", Nat. Protoc, 12, pp.255~278 (2017)
- Uen, Y. H., "Comparative proteomics, network analysis and post-translational modification identification reveal differential profiles of plasma Con A-bound glycoprotein biomarkers in gastric cancer", J. Proteom, 83, pp.197~213 (2013)
- Liebschner, D., "Polder maps : improving OMIT maps by excluding bulk solvent", Acta Crystallogr. D. Struct. Biol, 73, pp.148~157 (2017)
- Shinozaki, E., "Serum leucine-rich alpha-2-glycoprotein-1 with fucosylated triantennary N-glycan : a novel colorectal cancer marker", Bmc. Cancer, 18, pp.406~ (2018)
- Patwa, T. H., "Screening of glycosylation patterns in serum using natural glycoprotein microarrays and multi-lectin fluorescence detection", Anal. Chem, 78, pp.6411~6421 (2006)
- Hecht, M. J., "Neuropathy is a major contributing factor to diabetic erectile dysfunction", Neurol. Res, 23, pp.651~654 (2001)
- Musicki, B., "Endothelial dysfunction in diabetic erectile dysfunction", Int. J. Impot. Res, 19, pp.129~138 (2007)
- Rajagopal, B. S., "The hydrogen-peroxide-induced radical behaviour in human cytochrome c-phospholipid complexes : implications for the enhanced proapoptotic activity of the G41S mutant", Biochem. J, 456, pp.441~452 (2013)
- Ohtsubo, K., "Glycosylation in cellular mechanisms of health and disease", Cell, 126, pp.855~867 (2006)
- Varki, A., "Biological roles of glycans", Glycobiology, 27, pp.3~49 (2017)
- Vives, R. R., "Post-Synthetic Regulation of HS Structure : The Yin and Yang of the Sulfs in Cancer", Front. Oncol, 3, pp.331~ (2014)
- Reily, C., "Glycosylation in health and disease", Nat. Rev. Nephrol, 15, pp.346~366 (2019)
- Jackson, V. A., "Super-complexes of adhesion GPCRs and neural guidance receptors", Nat. Commun, 7, pp.11184~ (2016)
- Defeudis, G., "Erectile dysfunction and diabetes : A melting pot of circumstances and treatments", Diabetes Metab. Res. Rev, 38, pp.e3494~ (2022)
- Ashkenazy, H., "ConSurf 2016 : an improved methodology to estimate and visualize evolutionary conservation in macromolecules", Nucleic Acids Res, 44, pp.W344~W350 (2016)