PMC9682154_01
Clinical characteristics of the fetus in this study. (A) Ultrasound showed that the fetal cardiothoracic area ratio was increased, approximately 0.45, and the left atrium and left ventricle sizes were increased substantially. (B) Ultrasound showed hypertrophy of fetal myocardium and free fluid in the pericardial cavity (white arrow). (C) Ultrasound showed a large amount of free fluid in the fetal abdominal cavity and thickened abdominal skin tissue (white arrow). (D) After induced labor, autopsy revealed the heart was considerably enlarged. (E) Pathological sections showed thickening and fibrosis of the endocardial tissue (red arrows). (F) The muscular trabeculae were thick, the sinus recess of the myocardium persisted, the deep depression was staggered, the formation of dense myocardium in the corresponding area was reduced and thin, and the ventricular wall muscle layer remained loose (red circle).
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F1
In addition, 16 mm free fluid and thickening of abdominal subcutaneous tissue were seen in the abdominal cavity of the fetus (Figures 1A-C). The fetal heart tissue was taken for pathological examination, which showed that the endocardium was thickened and fibrotic, the dense myocardium was relatively thin, and the ventricular wall muscle layer remained loose (Figures 1D-F).
Information on the mutation found in this case. (A) The results of gene detection showed that TAFAZZIN had a missense mutation, c.311A > C/p.His104Pro. The father carried the wild-type form, while the mother carried the mutation. Sanger sequencing confirmed the existence of the mutation. (B) His104 is conserved between species (red box). (C) The crystal structure model analysis results show that the main chain N-H bond of His104 forms a hydrogen bond with Lys101, and the side chain forms a hydrogen bond with Glu73. When the mutation of His104Pro occurs, the hydrogen bond between Lys101 and Glu73 is broken because Pro104 has a five-membered ring structure with greater rigid strength than the straight-chain structure. Simultaneously, owing to the steric hindrance of Pro104, the secondary structure (part of the alpha-helix) may become a loop domain (red box). Therefore, the His104Pro mutation may reduce the stability of the local protein structure and affect its biological function.
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F2
Sanger sequencing confirmed the existence of the mutation (Figure 2A). In addition, because Pro104 is a five-membered ring containing an N structure with strong structural rigidity, the spatial steric resistance formed may cause the structure of the alpha-helix to become a loop, affecting its ability to interact with surrounding structures and resulting in reduced local structural stability (Figures 2B,C).
Western blotting results show that the protein expression in TAFAZZIN-MUT (the His104Pro mutant) is significantly lower than that in TAFAZZIN-WT (the wild type), indicating that His104Pro may reduce human TAFAZZIN protein expression and affect relevant biological functions. *, p < 0.05; NC, negative control.
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F3
The constructed plasmid was transfected into human Hek293T cells with 5 mul Lip2000 reagent (Thermo Fisher Scientific Inc., Waltham, MA, USA). The protein (80 mug) was extracted with protein lysate (Beyotime Inc., Shanghai, China), and the protein expression was examined by WB analysis. The results were analyzed using ImageJ software, and it was found that the protein expression of the TAFAZZIN-MUT group was significantly lower than that of the TAFAZZIN-WT group; it was reduced by approximately 31.37% (p < 0.001). This indicates that the variation in His104Pro might lead to a decrease in protein expression, thereby affecting protein function (Figure 3).