Discovery of the Recessive Effect of Human Polymerase δ Proofreading Deficiency: Through Mutation Analysis of Normal Cells and Cancer Cells with POLD1 Mutations

Background Introduction

The disruption of DNA repair is one of the main mechanisms leading to hereditary cancer. Heterozygous pathogenic variants in the exonic regions of POLD1 and POLE affect the proofreading activity of exonucleases, resulting in cancer susceptibility syndromes, manifesting as increased risks of gastrointestinal multiple polyps, colorectal cancer, endometrial cancer, etc. The current general explanation is that the loss of these exonuclease activities is linked to cancer development through increased somatic mutation rates. However, specific verification of this hypothesis remains controversial. Researchers explored this issue by analyzing fibroblast clones derived from members of a family carrying POLD1 mutations and de novo mutations between parents and children.

Paper Source

This study was conducted by Maria A. Andrianova, Vladimir B. Seplyarskiy, Mariona Terradas, and others from multiple research institutions including the Institute for Research in Biomedicine (IRB Barcelona), Harvard Medical School, and the Catalan Institute of Oncology. The research findings were published in the European Journal of Human Genetics in 2024.

Research Workflow

Researchers focused on a specific family with multiple members heterozygously carrying POLD1 pathogenic variants. The study first sequenced the genomes of 7 POLD1 L474P heterozygous carriers and 5 wild-type family members. They then analyzed the exomes or genomes of tumors developed by three carriers of different constitutional pathogenic POLD1 variants (L474P, D316H, and S478N). Using single-cell-derived fibroblast clones cultured for about 40 generations from POLD1 L474P positive and negative family members, they estimated the role of heterozygous POLD1 L474P in somatic mutation effects and detected de novo germline mutations through robust methods.

Main Research Findings

1. Heterozygous POLD1 L474P causes only a slight increase in mutation rates in somatic cells, but in related cancers, somatic inactivation of the POLD1 wild-type copy leads to extremely high mutation rates.
2. Somatic inactivation of the POLD1 wild-type allele is involved in tumor development. These results suggest that Polymerase δ proofreading deficiency has a recessive effect on mutation rates.
3. By comparing somatic fibroblast clones of family members and de novo mutations in children, it was found that POLD1 L474P only slightly increases mutation rates in both germline and somatic cells.

Research Conclusions and Significance

This study demonstrates the minimal impact of heterozygous POLD1 pathogenic variants on mutation rates in human somatic and germ cells, even though such heterozygous carriers tend to develop highly mutable or hypermutable cancers. We observed for the first time that the hypermutability in cancers or adenomas is associated with the somatic loss of the POLD1 wild-type allele. These results obtained in human cells and tumors correspond to the extensive literature discussing the recessive effect of POLD1 proofreading deficiency in yeast and mice.

Research Highlights

1. Novel research methods: Combining single-cell-derived fibroblast clone culture with genome sequencing of healthy family members.
2. Importance of findings: Revealing a new possible mechanism for cancer development, namely the dramatic increase in mutation rates caused by somatic inactivation of the POLD1 wild-type allele.
3. Guidance for future clinical work: The classification of POLD1 variants may need to consider tumor relevance and the appearance of the SBS10D mutation signature.