Germline Mutations in FAF1 Are Associated With Hereditary Colorectal Cancer.
Adaptor Proteins, Signal Transducing
/ genetics
Aged
Apoptosis
/ genetics
Apoptosis Regulatory Proteins
/ genetics
Cell Line, Tumor
Colorectal Neoplasms
/ genetics
DNA Mutational Analysis
Female
Gene Knockout Techniques
Genetic Predisposition to Disease
Germ-Line Mutation
Humans
Male
Middle Aged
Mutagenesis, Site-Directed
NF-kappa B
/ metabolism
Neoplastic Syndromes, Hereditary
/ genetics
Pedigree
Exome Sequencing
Wnt Signaling Pathway
/ genetics
beta Catenin
/ metabolism
Functional Genomics
Gene Editing
Programmed Cell Death
Wnt Signaling
Journal
Gastroenterology
ISSN: 1528-0012
Titre abrégé: Gastroenterology
Pays: United States
ID NLM: 0374630
Informations de publication
Date de publication:
07 2020
07 2020
Historique:
received:
30
08
2019
revised:
19
02
2020
accepted:
08
03
2020
pubmed:
18
3
2020
medline:
1
4
2021
entrez:
18
3
2020
Statut:
ppublish
Résumé
A significant proportion of colorectal cancer (CRC) cases have familial aggregation but little is known about the genetic factors that contribute to these cases. We performed an exhaustive functional characterization of genetic variants associated with familial CRC. We performed whole-exome sequencing analyses of 75 patients from 40 families with a history of CRC (including early-onset cases) of an unknown germline basis (discovery cohort). We also sequenced specific genes in DNA from an external replication cohort of 473 families, including 488 patients with colorectal tumors that had normal expression of mismatch repair proteins (validation cohort). We disrupted the Fas-associated factor 1 gene (FAF1) in DLD-1 CRC cells using CRISPR/Cas9 gene editing; some cells were transfected with plasmids that express FAF1 missense variants. Cells were analyzed by immunoblots, quantitative real-time polymerase chain reaction, and functional assays monitoring apoptosis, proliferation, and assays for Wnt signaling or nuclear factor (NF)-kappa-B activity. We identified predicted pathogenic variant in the FAF1 gene (c.1111G>A; p.Asp371Asn) in the discovery cohort; it was present in 4 patients of the same family. We identified a second variant in FAF1 in the validation cohort (c.254G>C; p.Arg85Pro). Both variants encoded unstable FAF1 proteins. Expression of these variants in CRC cells caused them to become resistant to apoptosis, accumulate beta-catenin in the cytoplasm, and translocate NF-kappa-B to the nucleus. In whole-exome sequencing analyses of patients from families with a history of CRC, we identified variants in FAF1 that associate with development of CRC. These variants encode unstable forms of FAF1 that increase resistance of CRC cells to apoptosis and increase activity of beta-catenin and NF-kappa-B.
Sections du résumé
BACKGROUND & AIMS
A significant proportion of colorectal cancer (CRC) cases have familial aggregation but little is known about the genetic factors that contribute to these cases. We performed an exhaustive functional characterization of genetic variants associated with familial CRC.
METHODS
We performed whole-exome sequencing analyses of 75 patients from 40 families with a history of CRC (including early-onset cases) of an unknown germline basis (discovery cohort). We also sequenced specific genes in DNA from an external replication cohort of 473 families, including 488 patients with colorectal tumors that had normal expression of mismatch repair proteins (validation cohort). We disrupted the Fas-associated factor 1 gene (FAF1) in DLD-1 CRC cells using CRISPR/Cas9 gene editing; some cells were transfected with plasmids that express FAF1 missense variants. Cells were analyzed by immunoblots, quantitative real-time polymerase chain reaction, and functional assays monitoring apoptosis, proliferation, and assays for Wnt signaling or nuclear factor (NF)-kappa-B activity.
RESULTS
We identified predicted pathogenic variant in the FAF1 gene (c.1111G>A; p.Asp371Asn) in the discovery cohort; it was present in 4 patients of the same family. We identified a second variant in FAF1 in the validation cohort (c.254G>C; p.Arg85Pro). Both variants encoded unstable FAF1 proteins. Expression of these variants in CRC cells caused them to become resistant to apoptosis, accumulate beta-catenin in the cytoplasm, and translocate NF-kappa-B to the nucleus.
CONCLUSIONS
In whole-exome sequencing analyses of patients from families with a history of CRC, we identified variants in FAF1 that associate with development of CRC. These variants encode unstable forms of FAF1 that increase resistance of CRC cells to apoptosis and increase activity of beta-catenin and NF-kappa-B.
Identifiants
pubmed: 32179092
pii: S0016-5085(20)30336-X
doi: 10.1053/j.gastro.2020.03.015
pii:
doi:
Substances chimiques
Adaptor Proteins, Signal Transducing
0
Apoptosis Regulatory Proteins
0
CTNNB1 protein, human
0
FAF1 protein, human
0
NF-kappa B
0
beta Catenin
0
Types de publication
Journal Article
Observational Study
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
227-240.e7Informations de copyright
Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.