Journal: Children

Article Title: Thirteen New Patients of PPP2R5D Gene Mutation and the Fine Profile of Genotype–Phenotype Correlation Unraveling the Pathogenic Mechanism Underlying Macrocephaly Phenotype

doi: 10.3390/children11080897

Figure Lengend Snippet: Protein expression levels of PPP2R5D and its interaction with the PPP2CA and PPP2R1A. ( A ) HA-tagged PPP2R5D WT/variants were transfected in 293T cells, protein expression was detected through Western blot. A 66 kDa band was detected in all missense variants. As for frameshift variant H436Mfs*3, a ~50 kDa band was detected and it was significantly weaker. ( B ) HA-tagged PPP2R5D WT/variants were purified through HA pull down, the interaction with endogenous PPP2CA and PPP2R1A were detected through Western blot. No band of PPP2CA and PPP2R1A was detected in W207R, D251H, D251Y and H463Mfs*3. A weaker band of PPP2CA and PPP2R1A was detected in E198K, E200K, L203P and Q211P.

Article Snippet: The primary antibodies are listed as below: PPP2R5D (Abcam, #ab188323, Cambridge, UK), Vinculin (Cell Signaling Technology, #13901), PPP2R1A (NOVUS, #NBP2-19907, St. Louis, MO, USA), and PPP2CA (Cell Signaling Technology, #2308).

Techniques: Expressing, Transfection, Western Blot, Variant Assay, Purification

Journal: Children

Article Title: Thirteen New Patients of PPP2R5D Gene Mutation and the Fine Profile of Genotype–Phenotype Correlation Unraveling the Pathogenic Mechanism Underlying Macrocephaly Phenotype

doi: 10.3390/children11080897

Figure Lengend Snippet: Pathogenicity prediction of PPP2R5D and analysis of residue characterization and phenotype. ( A ) Pathogenicity prediction of PPP2R5D by AM, the yellow bar represents conserved region 1, 2, and 3. Observed pathogenic variants were represented by red dots. ( B ) Protein 3D structure of PP2A holoenzyme. PPP2R5D protein was colored by scores predicted by AM, from green to red, low to high. PPP2R1A and PPP2CA were colored grey and darker gray. Amino acids affected by pathogenic missense variants of PPP2R5D gene were labeled on the protein. ( C ) Pathogenicity prediction by AM of PPP2CA and PPP2R1A. ( D ) Two-sided Fisher’s exact test of phenotypes and characterization of residues. Circles represent the OR, and the horizontal bars represent 95% CI. Red circles mean that the features in the y axis were enriched in patients with the phenotype, green means the opposite.

Article Snippet: The primary antibodies are listed as below: PPP2R5D (Abcam, #ab188323, Cambridge, UK), Vinculin (Cell Signaling Technology, #13901), PPP2R1A (NOVUS, #NBP2-19907, St. Louis, MO, USA), and PPP2CA (Cell Signaling Technology, #2308).

Techniques: Residue, Labeling