The cell surface hyaluronidase TMEM2 is essential for systemic hyaluronan catabolism and turnover
Yuki Tobisawa, Naoki Fujita, Hayato Yamamoto, Chikara Οhyama, Fumitoshi Irie, Yu Yamaguchi
Abstract
As a major component of the extracellular matrix, hyaluronan (HA) plays an important role in defining the biochemical and biophysical properties of tissues. In light of the extremely rapid turnover of HA and the impact of this turnover on HA biology, elucidating the molecular mechanisms underlying HA catabolism is key to understanding the in vivo functions of this unique polysaccharide. Here, we show that TMEM2, a recently identified cell surface hyaluronidase, plays an essential role in systemic HA turnover. Employing induced global Tmem2 knockout mice (Tmem2iKO), we determined the effects of Tmem2 ablation not only on the accumulation of HA in bodily fluids and organs, but also on the process of HA degradation in vivo. Within 3 weeks of tamoxifen-induced Tmem2 ablation, Tmem2iKO mice exhibit pronounced accumulation of HA in circulating blood and various organs, reaching levels as high as 40-fold above levels observed in control mice. Experiments using lymphatic and vascular injection of fluorescent HA tracers demonstrate that ongoing HA degradation in the lymphatic system and the liver is significantly impaired in Tmem2iKO mice. We also show that Tmem2 is strongly expressed in endothelial cells in the subcapsular sinus of lymph nodes and in the liver sinusoid, two primary sites implicated in systemic HA turnover. Our results establish TMEM2 as a physiologically relevant hyaluronidase with an essential role in systemic HA catabolism in vivo, acting primarily on the surface of endothelial cells in the lymph nodes and liver. As a major component of the extracellular matrix, hyaluronan (HA) plays an important role in defining the biochemical and biophysical properties of tissues. In light of the extremely rapid turnover of HA and the impact of this turnover on HA biology, elucidating the molecular mechanisms underlying HA catabolism is key to understanding the in vivo functions of this unique polysaccharide. Here, we show that TMEM2, a recently identified cell surface hyaluronidase, plays an essential role in systemic HA turnover. Employing induced global Tmem2 knockout mice (Tmem2iKO), we determined the effects of Tmem2 ablation not only on the accumulation of HA in bodily fluids and organs, but also on the process of HA degradation in vivo. Within 3 weeks of tamoxifen-induced Tmem2 ablation, Tmem2iKO mice exhibit pronounced accumulation of HA in circulating blood and various organs, reaching levels as high as 40-fold above levels observed in control mice. Experiments using lymphatic and vascular injection of fluorescent HA tracers demonstrate that ongoing HA degradation in the lymphatic system and the liver is significantly impaired in Tmem2iKO mice. We also show that Tmem2 is strongly expressed in endothelial cells in the subcapsular sinus of lymph nodes and in the liver sinusoid, two primary sites implicated in systemic HA turnover. Our results establish TMEM2 as a physiologically relevant hyaluronidase with an essential role in systemic HA catabolism in vivo, acting primarily on the surface of endothelial cells in the lymph nodes and liver. Hyaluronan (HA), a member of the family of glycosaminoglycans (GAGs), is a long unbranched polysaccharide (as along as 25,000 disaccharide units in length) with a molecular weight reaching as high as 107 Da. HA is a major component of the extracellular matrix (ECM) and is also present in biological fluids, such as blood and lymph (1Laurent T.C. Fraser J.R. Hyaluronan.FASEB J. 1992; 6: 2397-2404Crossref PubMed Scopus (2041) Google Scholar, 2Hascall V. Esko J.D. Hyaluronan.in: rd Varki A. Cummings R.D. Esko J.D. Stanley P. Hart G.W. Aebi M. Darvill A.G. Kinoshita T. Packer N.H. Prestegard J.H. Schnaar R.L. Seeberger P.H. Essentials of Glycobiology. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY2015: 197-206Google Scholar, 3Toole B.P. Hyaluronan: From extracellular glue to pericellular cue.Nat. Rev. Cancer. 2004; 4: 528-539Crossref PubMed Scopus (1603) Google Scholar). HA is extremely hydrophilic and occupies a large hydrodynamic volume in solution. These unique biochemical and biophysical properties of HA are accompanied and influenced by another unique feature of HA metabolism—an extremely rapid turnover. An estimated one-third of total body HA (∼15 g in a person with a 70 kg body weight) is turned over daily (4Fraser J.R. Laurent T.C. Turnover and metabolism of hyaluronan.Ciba Found. Symp. 1989; 143 (discussion 53-49, 281-285): 41-53PubMed Google Scholar), and the metabolic half-life of HA in skin is only 1–1.5 days (5Stern R. Devising a pathway for hyaluronan catabolism: Are we there yet?.Glycobiology. 2003; 13: 105R-115RCrossref PubMed Scopus (281) Google Scholar). This is in stark contrast to half-lives of other ECM molecules in tissues, many of which have half-lives measured in months or years (6Price R.G. Spiro R.G. Studies on the metabolism of the renal glomerular basement membrane. Turnover measurements in the rat with the use of radiolabeled amino acids.J. Biol. Chem. 1977; 252: 8597-8602Abstract Full Text PDF PubMed Google Scholar, 7Verzijl N. DeGroot J. Thorpe S.R. Bank R.A. Shaw J.N. Lyons T.J. Bijlsma J.W. Lafeber F.P. Baynes J.W. TeKoppele J.M. Effect of collagen turnover on the accumulation of advanced glycation end products.J. Biol. Chem. 2000; 275: 39027-39031Abstract Full Text Full Text PDF PubMed Scopus (638) Google Scholar, 8Sivan S.S. Tsitron E. Wachtel E. Roughley P.J. Sakkee N. van der Ham F. DeGroot J. Roberts S. Maroudas A. Aggrecan turnover in human intervertebral disc as determined by the racemization of aspartic acid.J. Biol. Chem. 2006; 281: 13009-13014Abstract Full Text Full Text PDF PubMed Scopus (71) Google Scholar). This rapid turnover suggests that cells must possess extremely robust machinery for HA degradation. Degradation of HA is thought to be a multistep process. The widely accepted, although not thoroughly tested, model proposes that HA in the extracellular space is first partially degraded into intermediate-sized HA fragments by an extracellular or cell surface hyaluronidase (9Stern R. Jedrzejas M.J. Hyaluronidases: Their genomics, structures, and mechanisms of action.Chem. Rev. 2006; PubMed Scopus Google Scholar). intermediate-sized HA are and degraded by in and These HA fragments are degraded by into which are for the of or for (1Laurent T.C. Fraser J.R. Hyaluronan.FASEB J. 1992; 6: 2397-2404Crossref PubMed Scopus (2041) Google Scholar). the systemic of HA degradation not in a cell of the HA the ECM of is and by the of HA into lymphatic and is into the lymphatic the of degradation J.R. Laurent T.C. N. and degradation of hyaluronan in lymphatic J. PubMed Scopus Google Scholar). In HA present in blood is primarily and degraded by the liver (4Fraser J.R. Laurent T.C. Turnover and metabolism of hyaluronan.Ciba Found. Symp. 1989; 143 (discussion 53-49, 281-285): 41-53PubMed Google Scholar). major are to in HA degradation in tissues, as in Hyaluronan also as and and TMEM2 also as F. in hyaluronan catabolism Biol. PubMed Scopus Google Scholar). high levels of and a family are thought to have by R. The in the human and Biol. PubMed Scopus Google Scholar). the first identified member of the is expressed in the J.W. The functions of and Biol. PubMed Scopus Google Scholar). human as a T. R. and of human PubMed Scopus Google Scholar), identified by on with a human expressed in many a hyaluronidase with a of Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). a the of the identified in of cells a human expressed in many a hyaluronidase with a of Biol. Chem. Full Text Full Text PDF PubMed Scopus Google and is present in the of other cell and of human in and cell 2006; Full Text Full Text PDF PubMed Scopus Google Scholar, is in and other cell 2006; Full Text Full Text PDF PubMed Scopus Google Scholar), also to be on the cell surface a V. A. is a for the of which S. A. PubMed Scopus Google Scholar, J. M. is a PubMed Scopus Google Scholar). the of is present in the a of cells R.D. The as a and 2006; PubMed Scopus Google Scholar), also be on the cell surface a J.W. The functions of and Biol. PubMed Scopus Google Scholar). The for the hyaluronidase of and are a human expressed in many a hyaluronidase with a of Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, M. M. R. and of human PubMed Scopus Google Scholar), with the that primarily in the and P. J.W. The in of two hyaluronidase Biol. PubMed Scopus Google Scholar). family not only HA but also and (9Stern R. Jedrzejas M.J. Hyaluronidases: Their genomics, structures, and mechanisms of action.Chem. Rev. 2006; PubMed Scopus Google Scholar, S. of in the catabolism of Biol. PubMed Scopus Google Scholar). and TMEM2 a family of of the identified as a that HA A. A. M. T. S. S. a of is a hyaluronan in hyaluronan S. A. PubMed Scopus Google Scholar). with cells with the to of the HA of A. A. M. T. S. S. a of is a hyaluronan in hyaluronan S. A. PubMed Scopus Google Scholar), that plays a role in HA degradation. of cells HA A. A. M. T. S. S. a of is a hyaluronan in hyaluronan S. A. PubMed Scopus Google Scholar, A. P. A. S. J. the role of a in cell PubMed Scopus Google Scholar). HA degradation of the pathway A. A. M. T. S. S. a of is a hyaluronan in hyaluronan S. A. PubMed Scopus Google Scholar), that not be a hyaluronidase TMEM2 identified as of human that are only on the of T. M. A. R. E. J. J. J. of with as of Cancer. PubMed Scopus Google Scholar). The that TMEM2 with and that exhibit HA accumulation in S. J. is to and PubMed Scopus Google Scholar), to that TMEM2 be an cell surface we that TMEM2 is a that hyaluronidase T. F. of the is the cell surface Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). we that TMEM2 HA in a the that as a cell surface hyaluronidase F. A. The cell surface hyaluronidase TMEM2 cell and degradation of hyaluronan Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). TMEM2 is expressed widely in tissues, the lymph nodes and liver T. F. of the is the cell surface Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar), the primarily implicated in systemic HA In this we use Tmem2 knockout mice to the of TMEM2 in systemic HA to a role for TMEM2 in HA we have induced global Tmem2 ablation in mice. These knockout mice have to demonstrate a rapid accumulation of HA in bodily fluids and a of the of Tmem2 ablation, as as effects of Tmem2 ablation on ongoing degradation of HA in the vascular and lymphatic we show that TMEM2 is expressed in the endothelial cells in the lymph nodes and two sites implicated in systemic HA turnover. results establish that TMEM2 is a hyaluronidase that plays an essential role in systemic HA catabolism in vivo. the of TMEM2 in systemic HA we Tmem2 knockout mice using a Tmem2 and the is S. in by a of for in the Biol. PubMed Scopus Google Scholar). 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Hyaluronan.FASEB J. 1992; 6: 2397-2404Crossref PubMed Scopus (2041) Google Scholar, J.R. Laurent T.C. N. and degradation of hyaluronan in lymphatic J. PubMed Scopus Google Scholar). The rapid accumulation of HA in blood and lymph nodes observed in Tmem2iKO mice is with the that TMEM2 plays a physiologically relevant role in systemic HA We the in to Tmem2 in the lymph nodes and liver. In lymph of Tmem2 is primarily sites to and Tmem2 is robust in cells of the subcapsular sinus are in the lymphatic endothelial cells have implicated as the cell primarily for HA degradation in the lymphatic system J.R. Laurent T.C. N. and degradation of hyaluronan in lymphatic J. PubMed Scopus Google Scholar, The the hyaluronan 2003; 13: PubMed Scopus Google Scholar), we cell to endothelial cells and cells lymph nodes and Tmem2 by As in cells Tmem2 strongly with the results demonstrate that endothelial cells in lymph in in the subcapsular are the primary of Tmem2 In the Tmem2 is in cells that are along the liver sinusoid, exhibit Tmem2 This of suggests that Tmem2 is expressed in the liver endothelial This is with a J. S. R. M. S. V. of liver and cell Full Text Full Text PDF PubMed Scopus Google in the of liver and cell which Tmem2 in endothelial Tmem2 in liver endothelial we an of this show that Tmem2 are with endothelial cells the liver Our results HA accumulation in the of Tmem2 ablation strongly the of TMEM2 in systemic HA the accumulation of HA is an of HA catabolism and not establish the of a hyaluronidase in the ongoing process of HA degradation. we in vivo injection of HA model of hyaluronan turnover in the Biol. 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