Reduced expression of APC-1B but not APC-1A by the deletion of promoter 1B is responsible for familial adenomatous polyposis.

Kiyoshi Yamaguchi,Masami Arai,Seiya Imoto,Masashi Ueno,Yoichi Furukawa,Tsuneo Ikenoue,Satoshi Nagayama,Satoru Miyano,Tetsuo Shibuya,Seira Hatakeyama,Rui Yamaguchi,Eigo Shimizu,Mitsuhiro Komura

doi:10.1038/srep26011

Kiyoshi Yamaguchi, Masami Arai + Show 11 more

Open Access

https://doi.org/10.1038/srep26011

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Abstract

Germline mutations in the tumor suppressor gene APC are associated with familial adenomatous polyposis (FAP). Here we applied whole-genome sequencing (WGS) to the DNA of a sporadic FAP patient in which we did not find any pathological APC mutations by direct sequencing. WGS identified a promoter deletion of approximately 10 kb encompassing promoter 1B and exon1B of APC. Additional allele-specific expression analysis by deep cDNA sequencing revealed that the deletion reduced the expression of the mutated APC allele to as low as 11.2% in the total APC transcripts, suggesting that the residual mutant transcripts were driven by other promoter(s). Furthermore, cap analysis of gene expression (CAGE) demonstrated that the deleted promoter 1B region is responsible for the great majority of APC transcription in many tissues except the brain. The deletion decreased the transcripts of APC-1B to 39–45% in the patient compared to the healthy controls, but it did not decrease those of APC-1A. Different deletions including promoter 1B have been reported in FAP patients. Taken together, our results strengthen the evidence that analysis of structural variations in promoter 1B should be considered for the FAP patients whose pathological mutations are not identified by conventional direct sequencing.

Highlights

Familial adenomatous polyposis (FAP) is an autosomal dominant inherited disorder characterized by the development of hundreds to thousands of colorectal adenomas and subsequent progression to adenocarcinoma
Multiplex ligation-dependent probe amplification (MLPA) and amplicon sequencing using a panel of disease-associated genes were available for the detection of structural alteration of adenomatous polyposis coli (APC) and genetic alterations in other responsible genes, respectively, we took advantage of whole-genome sequencing (WGS) to test its potential in clinical genetic testing
We searched for deleterious variants in the genes associated with adenomatous polyposis of the colon (APC, MUTYH, NTHL1, POLD1, and POLE)

Summary

Introduction

Familial adenomatous polyposis (FAP) is an autosomal dominant inherited disorder characterized by the development of hundreds to thousands of colorectal adenomas and subsequent progression to adenocarcinoma. Recent studies have demonstrated that large deletions of the transcription regulatory region of APC result in reduced APC transcripts, and that they are involved in a small subset of FAP4,10–14 The latter cases include patients with biallelic germ line mutations in the base excision repair (BER) gene MUTYH, an autosomal-recessively inherited disease termed MUTYH-associated polyposis (MAP)[15]. We have applied whole-genome sequencing (WGS) for a patient with both multiple adenomatous polyps in the colon and advanced rectal cancer, and without family history of polyposis This analysis identified a de novo deletion in the promoter region of APC, which markedly decreased the transcription of the mutant allele. These data indicate that NGS improves genetic diagnosis of hereditary diseases whose mutations have been overlooked by routine genetic testing, and that analysis of allele specific expression may help the determination of pathological changes in the promoter region of responsible genes

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