Identification and Functional Evaluation of a Novel TBX4 Mutation Underlies Small Patella Syndrome

Abstract

Small patella syndrome (SPS) is a rare autosomal dominant disorder caused by mutations in TBX4 gene which encodes a transcription factor of FGF10. However, how TBX4 mutations result in SPS is poorly understood. Here, a novel TBX4 mutation c.1241C>T (p.P414L) was identified in a SPS family and series of studies were performed to evaluate the influences of TBX4 mutations (including c.1241C>T and two known mutations c.256G>C and c.743G>T). Results showed that mesenchymal stem cells (MSCs) with stable overexpression of either TBX4 wild-type (TBX4wt) or mutants (TBX4mt) were successfully generated. Immunofluorescence study revealed that both the overexpressed TBX4 wild-type and mutants were evenly expressed in the nucleus suggesting that these mutations do not alter the translocation of TBX4 into the nucleus. Interestingly, MSCs overexpression of TBX4mt exhibited reduced differentiation activities and decreased FGF10 expression. Chromatin immunoprecipitation (ChIP) study demonstrated that TBX4 mutants still could bind to the promoter of FGF10. However, dual luciferase reporter assay clarified that the binding efficiencies of TBX4 mutants to FGF10 promoter were reduced. Taken together, MSCs were firstly used to study the function of TBX4 mutations in this study and the results indicate that the reduced binding efficiencies of TBX4 mutants (TBX4mt) to the promoter of FGF10 result in the abnormal biological processes which provide important information for the pathogenesis of SPS.

Long-Term Effect of TBX4 Germline Mutation on Pulmonary Clinico-Histopathologic Phenotype

“…In neonatal-onset PAH cases with TBX4 variants pulmonary growth arrest is demonstrated by acinar dysplasia and simplified alveolar development. 13,17,22,25 In addition to disrupted respiratory tract development and hindlimb hypoplasia, neurodevelopmental disorders and dysmorphic facial features are common in patients with TBX4 variants. It is hypothesized that TBX4 has a supplemental role in the formation of external genitalia, anorectum, and body wall.…”

“…Tbx4 protein has been shown to regulate expression of fibroblast growth factor 10 (FGF10), as well as activation of the Wnt pathway to induce fetal lung branching and lobe separation, as well as chondrogenesis and limb bud outgrowth. 8,12,13,17 TBX5, a neighboring T-box family gene, has an analogous role in upper limb development but has no identified role in lung development or longterm pulmonary health. 8,13 The complex pathobiology of the Tbx4-PAH pathway allows for multiple potential therapeutic targets, though there are currently no drugs available on the market that provide a cure for PAH.…”

“…In neonatal-onset PAH cases with TBX4 variants pulmonary growth arrest is demonstrated by acinar dysplasia and simplified alveolar development. 13,17,22,25…”

“…[3][4][5][7][8][9][10] Tbx4 protein, expressed embryologically in developing lung mesenchyme and lower extremity limb buds, contributes to distal lung development and hind-limb formation. [11][12][13][14][15][16][17][18][19][20] Past studies have shown a bimodal age distribution of the onset of clinical disease in TBX4-related PAH: 1) neonatal patients with severe and often lethal pulmonary hypertension with respiratory failure; and 2) PAH associated with variable severity of chronic lung disease, which can present late in childhood and in adults. 7,8,9,[21][22][23][24] Structural features of TBX4-related PAH and associated lung disease, especially in subjects presenting beyond the immediate postnatal period, have been incompletely characterized.…”