TENT5A-associated osteogenesis imperfecta: long-term follow-up and molecular insights

Abstract

OI is a genetically diverse disorder characterized by bone fragility and deformities, with most cases attributed to dominant mutations in collagen I-related genes. However, mutations in TENT5A, encoding a poly(A) polymerase, have recently been implicated in severe, recessive forms of OI. We reported 3 individuals from a consanguineous family with a novel homozygous TENT5A variant (c.672G>T, p.Arg224Ser). Our patients presented with severe bone fragility, generalized osteopenia, skeletal deformities, short stature, and early wheelchair dependence. Patient-derived fibroblasts demonstrated impaired collagen secretion and disorganized fibril network formation. Our findings provide new insights into the complex pathophysiology of TENT5A-associated OI and underscore the need for further research into its role in skeletal homeostasis.

Keywords: FAM46a; bisphosphonates; collagen I; growth hormone deficiency; skeletal dysplasias.

Figure 1

X-ray images of the 3 affected patients. Each panel is labeled with the respective patient. (A) Pedigree of the family from the affected individuals. (B) Whole body DXA scan of patient PII-1 showing severe skeletal deformities, generalized osteopenia, and severe obesity. (C-E) Lateral spine X-rays of all 3 patients at the age of 22 yr, 14 yr, and 3 yr showing vertebral compressions in all patients. (F) Right tibia of patient PII-1 at the age of 8 yr, showing osteopenia and severe deformity. (G) Left tibia of patient PII-2 at the age of 8 yr. (H) Right femur at the age of 3 yr from patient PIV-1. Note the generalized osteopenia, severe bowing, and thin cortex with horizontal bright lines from bisphosphonate therapy deposits. (I) Thoracic X-ray at the age of 20 yr, highlighting thin and hypertransparent ribs. Created with https://www.biorender.com. For visualization, all images were adjusted in brightness.

Figure 2

Cellular consequences of the (c.672G>T) variant in TENT5A leading to an amino acid exchange at the protein level (p.Arg224Ser). (A) Predicted protein structure indicating the localization of the mutation (p.Arg224Ser) in TENT5A. (B) Relative gene expression of TENT5A, COL1A1 and COL1A2 in control and patient (P) fibroblasts at passages 4-12 was assessed by qPCR. Gene expression was normalized to GAPDH, calibrated to the mean of controls and fold changes are plotted from n = 3 independent experiments. Statistical analysis: one-way ANOVA with Bonferroni’s multiple comparison test; # statistically different to all other samples (p ≤ .05), § statistically different to first control sample (p ≤ .05). (C) Immunoblot analysis of collagen type I from the cell layer of control and patient (P) fibroblast cultures at passages 3-12. GAPDH was used as loading control. Size of marker bands is indicated. Band intensities were quantified using ImageJ analysis software, normalized to GAPDH and calibrated to mean of all controls (n = 3 independent experiments). (D) Immunoblot analysis of collagen type I from the cell culture media of control and patient (P) fibroblast cultures at passages 3-12. Ponceau staining was used as loading control. Size of marker bands is indicated. Note that multiple COL I bands are detected with the upper band being more abundant in controls while in patient cells the lower band dominates (arrowheads, enlarged for better visibility). Statistical analysis: one-way ANOVA with Bonferroni’s multiple comparison test; # statistically different to first control and patient sample (p ≤ .05). (E) Fluorescence staining using collagen specific antibodies (COL I) and collagen hybridizing peptide (CHP) simultaneously to detect collagen-I in control and patient (P) fibroblast cultures at passages 5-11. Intracellular detection of collagen are highlighted (open arrowheads). For visualization, images were adjusted in brightness. Scale bar: 50 μm for all images in (E). (F) Negative staining and transmission electron microscopy of collagen fibrils secreted into cell culture media from control and patient (P) cells at passages 4-10. Scale bar: 100 nm for all images in (F).