Collagen IVα345 dysfunction in glomerular basement membrane diseases. I. Discovery of a COL4A3 variant in familial Goodpasture’s and Alport diseases

Abstract

Diseases of the glomerular basement membrane (GBM), such as Goodpasture’s disease (GP) and Alport syndrome (AS), are a major cause of chronic kidney failure and an unmet medical need. Collagen IVα345 is an important architectural element of the GBM that was discovered in previous research on GP and AS. How this collagen enables GBM to function as a permselective filter and how structural defects cause renal failure remain an enigma. We found a distinctive genetic variant of collagen IVα345 in both a familial GP case and four AS kindreds that provided insights into these mechanisms. The variant is an 8-residue appendage at the C-terminus of the α3 subunit of the α345 hexamer. A knock-in mouse harboring the variant displayed GBM abnormalities and proteinuria. This pathology phenocopied AS, which pinpointed the α345 hexamer as a focal point in GBM function and dysfunction. Crystallography and assembly studies revealed underlying hexamer mechanisms, as described in Boudko et al. and Pedchenko et al. Bioactive sites on the hexamer surface were identified where pathogenic pathways of GP and AS converge and, potentially, that of diabetic nephropathy (DN). We conclude that the hexamer functions include signaling and organizing macromolecular complexes, which enable GBM assembly and function. Therapeutic modulation or replacement of α345 hexamer could therefore be a potential treatment for GBM diseases, and this knock-in mouse model is suitable for developing gene therapies. Diseases of the glomerular basement membrane (GBM), such as Goodpasture’s disease (GP) and Alport syndrome (AS), are a major cause of chronic kidney failure and an unmet medical need. Collagen IVα345 is an important architectural element of the GBM that was discovered in previous research on GP and AS. How this collagen enables GBM to function as a permselective filter and how structural defects cause renal failure remain an enigma. We found a distinctive genetic variant of collagen IVα345 in both a familial GP case and four AS kindreds that provided insights into these mechanisms. The variant is an 8-residue appendage at the C-terminus of the α3 subunit of the α345 hexamer. A knock-in mouse harboring the variant displayed GBM abnormalities and proteinuria. This pathology phenocopied AS, which pinpointed the α345 hexamer as a focal point in GBM function and dysfunction. Crystallography and assembly studies revealed underlying hexamer mechanisms, as described in Boudko et al. and Pedchenko et al. Bioactive sites on the hexamer surface were identified where pathogenic pathways of GP and AS converge and, potentially, that of diabetic nephropathy (DN). We conclude that the hexamer functions include signaling and organizing macromolecular complexes, which enable GBM assembly and function. Therapeutic modulation or replacement of α345 hexamer could therefore be a potential treatment for GBM diseases, and this knock-in mouse model is suitable for developing gene therapies. Diseases of the glomerular basement membrane (GBM) are a major cause of chronic kidney disease, a health problem affecting about 10% of the global population and an unmet medical need (1Levin A. Tonelli M. Bonventre J. Coresh J. Donner J.A. Fogo A.B. Fox C.S. Gansevoort R.T. Heerspink H.J.L. Jardine M. Kasiske B. Kottgen A. Kretzler M. Levey A.S. Luyckx V.A. et al.Global kidney health 2017 and beyond: A roadmap for closing gaps in care, research, and policy.Lancet. 2017; 390: 1888-1917Abstract Full Text Full Text PDF PubMed Scopus (370) Google Scholar, 2Bikbov B. Perico N. 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Med. 2003; 348: 2543-2556Crossref PubMed Scopus (697) Google Scholar, 5Naylor R.W. Morais M. Lennon R. Complexities of the glomerular basement membrane.Nat. Rev. Nephrol. 2021; 17: 112-127Crossref PubMed Scopus (12) Google Scholar). These abnormalities involve structural alterations in collagen IV, the major GBM component. How collagen IV enables GBM to function as a permselective filter and how its structural alterations cause GBM abnormalities and dysfunction remain an enigma. New insights into these mysteries were revealed in the present study of a distinctive genetic variant of collagen IV, uniquely associated with both AS and GP diseases, coupled with crystallography and animal studies. Collagen IV is a family of six homologous α-chains, which are distributed within basement membranes (BMs) that underlie the epithelial architecture of the nephron. In pioneering studies of the GBM over 50 years ago, collagen IV was identified as a novel collagen and shown to be structurally altered in DN (6Kefalides N.A. Isolation and characterization of the collagen from glomerular basement membrane.Biochemistry. 1968; 7: 3103-3112Crossref PubMed Scopus (110) Google Scholar, 7Kefalides N.A. Structure and biosynthesis of basement membranes.Int. Rev. Connect. Tissue Res. 1973; 6: 63-104Crossref PubMed Google Scholar, 8Spiro R.G. Studies on the renal glomerular basement membrane. Preparation and chemical composition.J. Biol. Chem. 1967; 242: 1915-1922Abstract Full Text PDF PubMed Google Scholar, 9Beisswenger P.G. Spiro R.G. Human glomerular basement membrane: Chemical alteration in diabetes mellitus.Science. 1970; 168: 596-598Crossref PubMed Scopus (116) Google Scholar, 10Hudson B.G. Spiro R.G. Studies on the native and reduced alkylated renal glomerular basement membrane. Solubility, subunit size, and reaction with cyanogen bromide.J. Biol. 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N. basement membrane disease in human Nephrol. 2017; PubMed Scopus Google the variant was found in a population of of with health and in a of of the in an of about harboring the variant The was identified in six with renal These were identified in a of patients to of and as of genetic syndrome, focal and or of genetic renal diseases. In was in in as of the syndrome in as of the in as of the Alport and in was for genetic renal diseases. a of these patients a In the was identified in a of six from four and of with renal ranging from or to kidney disease and on the of in or J. G. K. K. A. M. Alport A of genetic of collagen IV A of the Alport Int. 2018; Full Text Full Text PDF PubMed Scopus Google these six described were as AS. 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Hudson B.G. et from by 2017; PubMed Scopus Google Scholar, R.G. The in Goodpasture's A model for to Int. Full Text Full Text PDF PubMed Scopus Google Scholar, H. Hudson B.G. The and in Goodpasture's 2017; PubMed Scopus Google on a α345 hexamer model J. K. A. Hudson B.G. Molecular in the assembly of are in the of triple helical Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The of these pathogenic pathways that a on the surface of the α345 hexamer is in of GP and the is a into the GBM, and its on hexamer function as a for the of on the hexamer these to the of the in a mouse model and to the and by the structure and assembly of the α345 hexamer. a mouse harboring the variant the within of the gene were of a and in an as a A of a and and was shown in were to with the a for and and the were and to for four to on were and was and were and in the additional on mouse and knock-in on a were and in the The of collagen IVα345 to the GBM by the was in the and was that the into the was from the of the α3 from mouse and of kidney revealed with in The GBM was in with and with revealed and GBM thickening The of from to and displayed a in at to in the appendage antibodies were displayed GBM and This is from the which the J.A. R. W.J. G.C. Collagen A mouse model for Alport PubMed Scopus Google Scholar, Molecular and defects in of collagen for Alport Cell Biol. PubMed Scopus Google Scholar). The the of the in the function of the collagen IVα345 these revealed that the the α345 hexamer a of abnormalities in the GBM, and with of displayed GBM for AS GBM The in the the in and that α345 hexamer the In to mouse that the collagen IVα345 from basement the mouse to functions of the α345 at a specific and to pathogenic mechanisms. The of that the appendage is a in GP The in both and pinpointed the α345 hexamer as a the GBM to function as a permselective In the present study of diseases, familial GP and AS, a genetic collagen IVα345 variant a 8-residue to both diseases, was found to as a into the of the The appendage is pathogenic in abnormalities in the GBM, and These phenocopied the of glomerular in human AS, which the of and as AS J. G. K. K. A. M. Alport A of genetic of collagen IV A of the Alport Int. 2018; Full Text Full Text PDF PubMed Scopus Google Scholar). the prevalence of the is to be at of to renal failure and The structure and assembly of the α345 as in Boudko et al. R. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. structure of the α345 Biol. Chem. 2021; Full Text Full Text PDF Scopus Google and Pedchenko et al. V. Boudko M. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. A for α345 hexamer Biol. Chem. 2021; Full Text Full Text PDF Scopus Google provided a to how the a AS a role in AS and a structural for GP R. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. structure of the α345 Biol. Chem. 2021; Full Text Full Text PDF Scopus Google Scholar, V. Boudko M. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. A for α345 hexamer Biol. Chem. 2021; Full Text Full Text PDF Scopus Google Scholar). a with This in in with or and with and Boudko et al. R. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. structure of the α345 Biol. Chem. 2021; Full Text Full Text PDF Scopus Google of Google Scholar, and for Rev. PubMed Scopus Google Scholar). The appendage is with the GP of the R. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. structure of the α345 Biol. Chem. 2021; Full Text Full Text PDF Scopus Google Scholar). This of pathogenic pathways that the is functions that include signaling and organizing macromolecular R. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. structure of the α345 Biol. Chem. 2021; Full Text Full Text PDF Scopus Google Scholar, V. Boudko M. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. A for α345 hexamer Biol. Chem. 2021; Full Text Full Text PDF Scopus Google Scholar). the is V. Boudko M. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. A for α345 hexamer Biol. Chem. 2021; Full Text Full Text PDF Scopus Google its functions be by the and genetic that in the hexamer in AS and in GP and in DN and Boudko et al. R. J. Hudson B.G. Collagen IVα345 dysfunction in glomerular basement membrane diseases. structure of the α345 Biol. Chem. 2021; Full Text Full Text PDF Scopus Google Scholar). conclude that the α345 hexamer is a focal point with GBM and its function as a permselective which be altered in GBM α345 hexamer is a focal point in GBM function and dysfunction with sites as potential in the treatment of GBM kidney diseases. The collagen IVα345 is a major structure underlying glomerular (GBM) basement membrane. In the GBM, is by of the glomerular in the Nephrol. Full Text Full Text PDF PubMed Scopus Google in the of protomers that in the of of C.F. Pedchenko V. M. M. A.S. R. G. et collagen IV network assembly basement membrane Cell Biol. PubMed Scopus Google Scholar). The as a C.F. Hudson B.G. collagen IV on the of 2017; PubMed Scopus Google due to the of sites and that in and M. chemical that and structure in and Rev. PubMed Scopus Google within the macromolecular of the basement membrane. The α345 hexamer is a within the collagen IVα345 The of the hexamer its specific and assembly by and the hexamer surface include within the basement membrane and surface are sites at the of the structure by where pathogenic for Alport syndrome, Goodpasture’s disease, and diabetic nephropathy This of pathogenic pathways that the signaling and organizing macromolecular complexes, which underlie the GBM The sites are for genetic and basement membrane abnormalities and to and are as both and in The sites and pathways are potential for of replacement and therapies. additional Boudko et al. R. J. Hudson B.G. 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