Identification of Novel PKD1 Mutations in Two Chinese Families with Autosomal Dominant Polycystic Kidney Disease by Targeted Next Generation Sequencing
Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common inherited kidney disorders, with an estimated incidence ranging from 1 in 400 to 1 in 1000 individuals. It is a late-onset systemic disorder characterized by the development and progressive enlargement of cysts in the kidneys, which ultimately leads to end-stage renal disease (ESRD). ADPKD is a heterogeneous monogenic disorder primarily caused by mutations in two genes: PKD1 and PKD2. Clinical data indicate that mutations in PKD1 account for approximately 85% of ADPKD cases, while PKD2 mutations are responsible for the remaining 15%. Notably, PKD1 mutations are associated with more severe disease manifestations, earlier onset, and a poorer prognosis compared to PKD2 mutations. Despite extensive research, no mutation hotspots have been identified for either gene, suggesting that mutations are highly variable and often unique to individual families.
In this study, two Chinese families with ADPKD were investigated using targeted next-generation sequencing (NGS) to identify mutations in the PKD1 gene. The study revealed a novel nonsense mutation, c.6491C>A (p.Ser2164Ter), and a known frameshift mutation, c.12608_12635del (p.Arg4203Profs*146), in the PKD1 gene. These findings contribute to the growing understanding of the genetic diversity of ADPKD and enrich the PKD1 mutation database, particularly in the Chinese population.
Clinical and Genetic Background of ADPKD
ADPKD is a systemic disorder that primarily affects the kidneys but can also involve other organs, including the liver, pancreas, and cardiovascular system. The disease is characterized by the formation of fluid-filled cysts in the renal tubules, which progressively enlarge and disrupt normal kidney function. Over time, this leads to a decline in renal function, culminating in ESRD, which necessitates dialysis or kidney transplantation. The clinical presentation of ADPKD is highly variable, with some individuals remaining asymptomatic until late adulthood, while others experience early-onset disease with rapid progression.
The genetic basis of ADPKD lies in mutations in the PKD1 and PKD2 genes, which encode the proteins polycystin-1 (PC1) and polycystin-2 (PC2), respectively. These proteins form a complex that localizes to the primary cilium of renal tubular epithelial cells, where they play a crucial role in mechanosensory functions, cell adhesion, proliferation, and differentiation. Disruption of this complex due to mutations in PKD1 or PKD2 leads to abnormal cellular signaling, hyperactive secretion, and the formation of cysts. The severity of the disease is often influenced by the specific mutation and its impact on the structure and function of the PC1-PC2 complex.
Study Design and Methodology
The study involved two three-generation Chinese families with a history of ADPKD. A total of 14 individuals from these families were recruited, all of whom had been definitively diagnosed with ADPKD using standard diagnostic criteria, including ultrasound, magnetic resonance imaging (MRI), or computed tomography (CT) scans. The clinical characteristics of the patients are summarized in Supplementary Table 1.
Targeted next-generation sequencing was employed to screen for mutations in the PKD1 gene. This approach allows for the simultaneous analysis of multiple genes or specific regions of interest with high accuracy and efficiency. The identified mutations were subsequently validated using Sanger sequencing, a gold standard method for confirming genetic variants.
Identification of Novel and Known PKD1 Mutations
In the first family (Pedigree 1), a novel nucleotide substitution, c.6491C>A, was identified in exon 15 of the PKD1 gene. This mutation was absent in 200 unrelated healthy individuals and had not been previously reported in any public genetic databases, including NCBI dbSNP, HapMap, Exome Variant Server Database, 1000 Genomes, and a database of 100 healthy Chinese adults. The variant was also absent in the Human Gene Mutation Database (HGMD), confirming its novelty. The c.6491C>A mutation was detected in all affected family members but was absent in asymptomatic individuals, demonstrating its co-segregation with the disease. Pathogenicity prediction using Mutation Taster classified the variant as “disease-causing.” The C to A transversion was predicted to introduce an early stop codon (UAA) at position 2164 (p.Ser2164Ter), resulting in a truncated protein that is 2140 amino acids shorter than the wild-type protein. This truncation would lead to the loss of the transmembrane domains and the cytoplasmic C-terminal tail of PC1, which are critical for its function.
In the second family (Pedigree 2), a known 28-base pair (bp) deletion, c.12608_12635del, was identified in exon 46 of the PKD1 gene. This mutation had been previously reported by Neumann et al. in the HGMD and is associated with ADPKD. The c.12608_12635del mutation also displayed co-segregation with the disease in the family. Pathogenicity prediction using Mutation Taster classified the variant as “disease-causing.” The 28-bp deletion was predicted to alter the open reading frame, resulting in the loss of the original stop codon (UAG) at position 4304 and the introduction of a new stop codon (UAA) at position 4348 (p.Arg4203Profs*146). This frameshift mutation would produce a prolonged protein with an abnormal cytoplasmic C-terminal tail, which is 44 amino acids longer than the wild-type protein. The abnormal C-terminal tail could disrupt normal signal transduction pathways, leading to pathogenic cystogenesis.
Functional Implications of the Identified Mutations
The PKD1 gene encodes polycystin-1 (PC1), a large membrane protein with an N-terminal extracellular region, eleven transmembrane domains, and a cytoplasmic C-terminal tail. The C-terminal tail contains a coiled-coil structure that interacts with polycystin-2 (PC2), forming the PC1-PC2 complex. This complex is essential for maintaining the structural and functional integrity of renal tubular epithelial cells. Mutations that disrupt the C-terminal tail of PC1 can impair its interaction with PC2, leading to abnormal cellular signaling and cyst formation.
In the case of the novel c.6491C>A mutation, the introduction of a premature stop codon would result in a truncated protein lacking the transmembrane domains and the cytoplasmic C-terminal tail. This would likely disrupt the PC1-PC2 complex, leading to defective mechanosensory functions and abnormal cell proliferation. Similarly, the known c.12608_12635del mutation would produce a protein with an abnormally long C-terminal tail, which could interfere with normal signal transduction and cellular processes. Both mutations would ultimately contribute to the hyperactive secretion and hyperplasia of tubular epithelial cells, resulting in the formation of renal cysts.
Conclusion
This study identified a novel nonsense mutation, c.6491C>A (p.Ser2164Ter), and a known frameshift mutation, c.12608_12635del (p.Arg4203Profs*146), in the PKD1 gene in two Chinese families with ADPKD. These findings highlight the genetic diversity of ADPKD and underscore the importance of mutation screening in different ethnic populations. The identification of novel mutations not only enriches the PKD1 mutation database but also provides valuable insights into the molecular mechanisms underlying ADPKD. Further research is needed to explore the functional consequences of these mutations and their potential implications for the development of targeted therapies for ADPKD.
doi.org/10.1097/CM9.0000000000000667
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