Mao Mao

Gould syndrome is caused by mutations in the genes encoding collagen IV alpha 1 (COL4A1) and 2 (COL4A2)—fundamental basement membrane components that form collagen α1α1α2(IV) heterotrimers

Basement membranes (BMs) are specialized extracellular matrix (ECM) structures essential for organ morphogenesis, architecture, and function. BM composition and properties vary between tissues, developmental stages, and disease states, and there is only a rudimentary understanding of BM dynamics. Here, we introduce a versatile mouse model carrying a multifunctional dual-color fluorescence tagged allele with knockout potential for the fundamental BM component type IV collagen alpha 1 (COL4A1). This allele enables the characterization of cell type- and time-specific contributions to BMs and the generation of a conditional Col4a1 null allele. We demonstrate the utility of this unique genetic resource in providing clinically relevant insights for individuals with Gould syndrome – a multisystem disorder caused by COL4A1 and COL4A2 mutations. We show active COL4A1 turnover in postnatal cerebrovascular BMs, identifying a potential interventional window for cerebrovascular manifestations associated with Gould syndrome. We also demonstrate that heterozygous Col4a1 deletion is significantly less pathogenic than dominant Col4a1 missense mutations, which has important implications for gene therapy.

Purpose: Mutations in the genes encoding type IV collagen alpha 1 (COL4A1) and alpha 2 (COL4A2) cause a multisystem disorder that includes ocular anterior segment dysgenesis (ASD) and glaucoma. We previously showed that transforming growth factor beta (TGFβ) signaling was elevated in developing anterior segments from Col4a1 mutant mice and that reducing TGFβ signaling ameliorated ASD, supporting a role for the TGFβ pathway in disease pathogenesis. Here, we tested whether altered TGFβ signaling also contributes to glaucoma-related phenotypes in Col4a1 mutant mice. Methods: To test the role of TGFβ signaling in glaucoma-relevant phenotypes, we genetically reduced TGFβ signaling using mice with mutated Tgfbr2, which encodes the common receptor for all TGFβ ligands in Col4a1^+/G1344D mice. We performed slit-lamp biomicroscopy and optical coherence tomography for qualitative and quantitative analyses of anterior and posterior ocular segments, histological analyses of ocular tissues and optic nerves, and intraocular pressure assessments using rebound tonometry. Results: Col4a1^+/G1344D mice showed defects of the ocular drainage structures, including iridocorneal adhesions, and phenotypes consistent with glaucomatous neurodegeneration, including thinning of the nerve fiber layer, retinal ganglion cell loss, optic nerve head excavation, and optic nerve degeneration. We found that reducing TGFβ receptor 2 (TGFBR2) was protective for ASD, ameliorated ocular drainage structure defects, and protected against glaucomatous neurodegeneration in Col4a1^+/G1344D mice. Conclusions: Our results suggest that elevated TGFβ signaling contributes to glaucomatous neurodegeneration in Col4a1 mutant mice.

Collagen type IV alpha 1 and alpha 2 (COL4A1 and COL4A2) are major components of almost all basement membranes. COL4A1 and COL4A2 mutations cause a multisystem disorder that can affect any organ but typically involves the cerebral vasculature, eyes, kidneys and skeletal muscles. In recent years, patient advocacy and family support groups have united under the name of Gould syndrome. The manifestations of Gould syndrome are highly variable, and animal studies suggest that allelic heterogeneity and genetic context contribute to the clinical variability. We previously characterized a mouse model of Gould syndrome caused by a Col4a1 mutation in which the severities of ocular anterior segment dysgenesis (ASD), myopathy and intracerebral hemorrhage (ICH) were dependent on genetic background. Here, we performed a genetic modifier screen to provide insight into the mechanisms contributing to Gould syndrome pathogenesis and identified a single locus [modifier of Gould syndrome 1 (MoGS1)] on Chromosome 1 that suppressed ASD. A separate screen showed that the same locus ameliorated myopathy. Interestingly, MoGS1 had no effect on ICH, suggesting that this phenotype could be mechanistically distinct. We refined the MoGS1 locus to a 4.3 Mb interval containing 18 protein-coding genes, including Fn1, which encodes the extracellular matrix component fibronectin 1. Molecular analysis showed that the MoGS1 locus increased Fn1 expression, raising the possibility that suppression is achieved through a compensatory extracellular mechanism. Furthermore, we found evidence of increased integrin-linked kinase levels and focal adhesion kinase phosphorylation in Col4a1 mutant mice that is partially restored by the MoGS1 locus, implicating the involvement of integrin signaling. Taken together, our results suggest that tissue-specific mechanistic heterogeneity contributes to the variable expressivity of Gould syndrome and that perturbations in integrin signaling may play a role in ocular and muscular manifestations.

Collagen type IV alpha 1 and alpha 2 (COL4A1 and COL4A2) are major components of almost all basement membranes. COL4A1 and COL4A2 mutations cause a multisystem disorder called Gould syndrome which can affect any organ but typically involves the cerebral vasculature, eyes, kidneys and skeletal muscles. The manifestations of Gould syndrome are highly variable and animal studies suggest that allelic heterogeneity and genetic context contribute to the clinical variability. We previously characterized a mouse model of Gould syndrome caused by a Col4a1 mutation in which the severities of ocular anterior segment dysgenesis (ASD), myopathy, and intracerebral hemorrhage (ICH) were dependent on genetic background. Here, we performed a genetic modifier screen to provide insight into the mechanisms contributing to Gould syndrome pathogenesis and identified a single locus ( modifier of Gould syndrome 1 ; MoGS1 ) on Chromosome 1 that suppressed ASD. A separate screen showed that the same locus ameliorated myopathy. Interestingly, MoGS1 had no effect on ICH, suggesting that this phenotype may be mechanistically distinct. We refined the MoGS1 locus to a 4.3 Mb interval containing 18 protein coding genes, including Fn1 which encodes the extracellular matrix component fibronectin 1. Molecular analysis showed that the MoGS1 locus increased Fn1 expression raising the possibility that suppression is achieved through a compensatory extracellular mechanism. Furthermore, we show evidence of increased integrin linked kinase levels and focal adhesion kinase phosphorylation in Col4a1 mutant mice that is partially restored by the MoGS1 locus implicating the involvement of integrin signaling. Taken together, our results suggest that tissue-specific mechanistic heterogeneity contributes to the variable expressivity of Gould syndrome and that perturbations in integrin signaling may play a role in ocular and muscular manifestations.