Validation and evolutionary conservation of the identified mutations in the patient with VUR and proteinuria.

<p>(A) Sanger sequencing analysis confirms the novel <i>COL4A3</i> mutations, <i>COL4A3_G/A_G695R</i> and <i>COL4A3_T/C_L1474P</i>, the <i>SALL2_G/C_G792R</i> and the <i>MYH9_C/T_L46F</i> mutations in the patient 1184405. (B) Multi species alignment shows that both the <i>COL4A3</i> mutation reference amino acids <i>Col4A3_ 695_G</i> and <i>Col4A3_1474_L</i>, the <i>SALL2_G792R</i> and the <i>MYH9_L46F</i> are highly conserved among species. (C) Schematic representation of Human <i>COL4A3</i> gene with the discovered mutations in patient 1184405. The domains are: signal peptide domain (red), the N terminal 7S domain (blue), the central triple helix collagenous (COL) domain (green) and the carboxy-terminal non-collagenous (NC1) domain (yellow).</p

Results of immunostaining with antibodies against α1, α3, and α5 chains of type IV collagen in the AS/VUR patient.

<p>(A) ColIVα1 expression is strong and consistent in the basement membranes (BM) of the glomeruli and the tubules; typically it is absent in adult glomeruli. Images show absent α3IV (B) and weak α5IV (C) staining in the glomerular BMs. (D&E) Electron micrograph from this patient shows segmental glomerular basement membrane (GBM) lamellation and outpouching (D arrow) and irregular thickening (E arrow) of the GBM characteristic of AS. COL4α3 immunostaining in control shows the normal tubular and capillary loop distribution for comparison (F).</p

Clinical characteristics of patients that underwent validation of custom multindexed targeted exome sequencing of 19 genes.

<p>Participants in italics are relatives in which the same mutation was found.</p

Pedigree analysis of the index patient with proetinuria and VUR.

<p>(A) Pedigree of the family of the index patient (1184405, arrow) shows presence of <i>Col4A3_T/C_L1474P</i> variant in one unaffected brother (4302442) and nephew (4302444), but absent in the unaffected sister (4302445) and niece (4302443); while the other damaging variant <i>Col4A3_G/A_G695R</i> is present only in the patient. The <i>SALL2</i> deleterious mutation <i>SALL2_G/C_G792R</i> is present in the patient and one unaffected brother (4302442) while the <i>MYH9_C/T_L46F</i> mutation is present in all but one affected family member (4302445). (B) Sanger sequencing analysis confirms the status of the novel <i>COL4A3</i> mutations <i>Col4A3_G/A_G695R</i> and <i>Col4A3_C/T_L1474P</i>, and the <i>SALL2_G/C_G792R</i> and <i>MYH9_C/T_L46F</i> mutations in the relatives of patient 1184405. The <i>Col4A3_T/C_L1474P</i> heterozygous variant was present in one unaffected brother (4302442) and nephew (4302444), but absent the unaffected sister (4302445) and niece (430443). The mutation <i>Col4A3_G/A_G695R</i> was absent in all other family members. The <i>SALL2_G/C_G792R</i> mutation was present only in the unaffected brother 4302442 while the <i>MYH9_C/T_L46F mutation</i> was more common and was present in everyone except the unaffected sister 4302445.</p