Cytoskeleton and nuclear lamina affection in recessive osteogenesis imperfecta: A functional proteomics perspective

Abstract

Osteogenesis imperfecta (OI) is a collagen-related disorder associated to dominant, recessive or X-linked transmission, mainly caused by mutations in type I collagen genes or in genes involved in type I collagen metabolism.Among the recessive forms, OI types VII, VIII, and IX are due to mutations in CRTAP, P3H1, and PPIB genes, respectively. They code for the three components of the endoplasmic reticulum complex that catalyzes 3-hydroxylation of type I collagen α1Pro986. Under-hydroxylation of this residue leads to collagen structural abnormalities and results in moderate to lethal OI phenotype, despite the exact molecular mechanisms are still not completely clear.To shed light on these recessive forms, primary fibroblasts from OI patients with mutations in CRTAP (n = 3), P3H1 (n = 3), PPIB (n = 1) genes and from controls (n = 4) were investigated by a functional proteomic approach. Cytoskeleton and nucleoskeleton asset, protein fate, and metabolism were delineated as mainly affected. While western blot experiments confirmed altered expression of lamin A/C and cofilin-1, immunofluorescence analysis using antibody against lamin A/C and phalloidin showed an aberrant organization of nucleus and cytoskeleton.This is the first report describing an altered organization of intracellular structural proteins in recessive OI and pointing them as possible novel target for OI treatment.SignificanceOI is a prototype for skeletal dysplasias. It is a highly heterogeneous collagen-related disorder with dominant, recessive and X-linked transmission. There is no definitive cure for this disease, thus a better understanding of the molecular basis of its pathophysiology is expected to contribute in identifying potential targets to develop new treatments.Based on this concept, we performed a functional proteomic study to delineate affected molecular pathways in primary fibroblasts from recessive OI patients, carrying mutations in CRTAP (OI type VII), P3H1 (OI type VIII), and PPIB (OI type IX) genes. Our analyses demonstrated the occurrence of an altered cytoskeleton and, for the first time in OI, of nuclear lamina organization. Hence, cytoskeleton and nucleoskeleton components may be considered as novel drug targets for clinical management of the disease.Finally, according to our analyses, OI emerged to share similar deregulated pathways and molecular aberrances, as previously described, with other rare disorders caused by different genetic defects.Those aberrances may provide common pharmacological targets to support classical clinical approach in treating different diseases.

The Osteocyte Transcriptome Is Extensively Dysregulated in Mouse Models of Osteogenesis Imperfecta

“…The results of our analysis confirm that in OI there is a significant dysregulation of transcripts involved in dendrite formation, ECM organization, collagen fibril organization, integrin‐mediated signaling pathway, tissue development, among others. This implies a profound alteration of cellular‐matrix‐tissue interactions that impact cellular function, morphology, and cytoskeletal organization and is consistent with recent evidence of cytoskeletal dysregulation in OI . Of the genes implicated in bone formation, transcripts involved in the WNT signaling pathway were differentially expressed in both our OI models.…”

“…This implies a profound alteration of cellularmatrix-tissue interactions that impact cellular function, morphology, and cytoskeletal organization and is consistent with recent evidence of cytoskeletal dysregulation in OI. (56,57) Of the genes implicated in bone formation, transcripts involved in the WNT signaling pathway were differentially expressed in both our OI models. These included at least 3 WNT ligands but also inhibitors of WNT signaling, suggesting a cellular attempt to increase osteoblast differentiation and function, perhaps triggered by the osteocyte sensing a defective matrix.…”

Genotypic and phenotypic characterization of Chinese patients with osteogenesis imperfecta

“…Variants in type I collagen ( n = 230) accounted for a majority of all cases (78% and 294/378 of all families, and 84% and 560/668 of all patients). Given that the ratios were slightly lower than previous estimations (Forlino & Marini, ; Gagliardi et al, ; Tournis & Dede, ), this deviation might be due to the trend for individuals with a severe phenotype to seek diagnosis and treatment, the existence of patients misdiagnosed with OI, or to potential causative genes that have not yet been identified.…”

“…The variants in COL1A1 / COL1A2 can be categorized into quantitative and structural defects (substitutions of glycine in most cases; Hald et al, ). Hitherto, at least 15 noncollagen genes ( IFITM5 , SERPINF1 , CRTAP , P3H1 , PPIB , SERPINH1 , FKBP10 , PLOD2 , BMP1 , SP7 , TMEM38B , WNT1 , CREB3L1 , SPARC, and MBTPS2 ) have been identified in OI patients (Byers & Pyott, ; Gagliardi et al, ; Lindert et al, ; Rohrbach & Giunta, ).…”

Mechanical property characterization of hundreds of single nuclei based on microfluidic constriction channel

“…Nuclear lamina consisting of lamins and nuclear lamin‐associated membrane proteins is a fibrillar network structure that exists in the nucleus of most eukaryotic cells . Such a fibrillar network structure of nuclear lamina connects the inner nuclear membrane inside a nucleus, and the endoplasmic reticulum in cytoplasm not only providing the mechanical support of nucleus , but also involving several cellular events [e.g., chromatin organization , cell cycle regulation , cell differentiation , DNA replication ]. Reports in literature have demonstrated that the variations of mechanical properties of nuclei are associated with physiological and pathological status of cells including tumor development , stem cell differentiation , and blood cell relevant diseases .…”