PUBLICATION

Mutations in RAD21 Disrupt Regulation of APOB in Patients with Chronic Intestinal Pseudo-obstruction

Authors

Bonora, E., Bianco, F., Cordeddu, L., Bamshad, M., Francescatto, L., Dowless, D., Stanghellini, V., Cogliandro, R.F., Lindberg, G., Mungan, Z., Cefle, K., Ozcelik, T., Palanduz, S., Ozturk, S., Gedikbasi, A., Gori, A., Pippucci, T., Graziano, C., Volta, U., Caio, G., Barbara, G., D'Amato, M., Seri, M., Katsanis, N., Romeo, G., De Giorgio, R.

ID

ZDB-PUB-150113-12

Date

2015

Source

Gastroenterology 148(4): 771-782.e11 (Journal)

Registered Authors

Francescatto, Ludmila, Katsanis, Nicholas

Keywords

animal model, genetic analysis, intestinal motility, sporadic and familial chronic intestinal pseudo-obstruction

MeSH Terms

Adult
Animals
Apolipoprotein B-100/genetics*
Case-Control Studies
Cell Cycle Proteins/genetics*
Chronic Disease
Core Binding Factor Alpha 2 Subunit/genetics*
Enteric Nervous System/metabolism*
Enteric Nervous System/physiopathology
Female
Gastrointestinal Motility/genetics*
Gene Expression Profiling
Gene Knockdown Techniques
HEK293 Cells
Humans
Intestinal Pseudo-Obstruction/genetics*
Intestinal Pseudo-Obstruction/physiopathology
Male
Middle Aged
Nuclear Proteins/genetics*
Phosphoproteins/genetics*
RNA, Messenger/genetics*
Sequence Analysis, DNA
Young Adult
Zebrafish
Zebrafish Proteins/genetics*

PubMed

25575569 Full text @ Gastroenterology

Abstract

Background & Aims: Chronic intestinal pseudo-obstruction (CIPO) is characterized by severe intestinal dysmotility that mimicks a mechanical sub-occlusion with no evidence of gut obstruction. We searched for genetic variants associated with CIPO to increase our understanding of its pathogenesis and indentify potential biomarkers.

Methods We performed whole-exome sequencing of genomic DNA from patients with familial CIPO syndrome. Blood and lymphoblastoid cells were collected from patients and controls (individuals without CIPO); levels of mRNA and proteins were analyzed by quantitative reverse transcription PCR, immunoblot, and mobility shift assays. cDNAs were transfected into HEK293 cells. Expression of rad21 was suppressed in zebrafish embryos using a splice-blocking morpholino (rad21a MO). Gut tissues were collected and analyzed.

Results We identified a homozygous mutation (p.622, encodes Ala>Thr) in RAD21 in patients from a consanguineous family with CIPO. Expression of RUNX1, a target of RAD21, was reduced in cells from patients with CIPO compared with controls. In zebrafish, suppression of rad21a reduced expression of runx1; this phenotype was corrected by injection of human RAD21 mRNA, but not with the mRNA from the mutated p.622 allele. rad21a MO zebrafish had delayed intestinal transit and greatly reduced numbers of enteric neurons, similar to patients with CIPO. This defect was greater in zebrafish with suppressed expression of ret and rad21, indicating their interaction in regulation of gut neurogenesis. The promoter region of APOB bound RAD21 but not RAD21 p.622 Ala>Thr; expression of wild-type RAD21 in HEK293 cells repressed expression of APOB, compared with control vector. The gut-specific isoform of APOB (APOB48) is overexpressed in sera from patients with CIPO who carry the RAD21 mutation. APOB48 is also overexpressed in sporadic CIPO in sera and gut biopsies.

Conclusions Some patients with CIPO carry mutations in RAD21 that disrupt the ability of its product to regulate genes such as RUNX1 and APOB. Reduced expression of rad21 in zebrafish, and dysregulation of these target genes, disrupts intestinal transit and development of enteric neurons.

Genes / Markers

Figures

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Expression

Phenotype

Mutations / Transgenics

Human Disease / Model

Sequence Targeting Reagents

Fish

Antibodies

Orthology

Engineered Foreign Genes

Mapping

Bonora et al., 2015 ZDB-PUB-150113-12 PMID:25575569

Mutations in RAD21 Disrupt Regulation of APOB in Patients with Chronic Intestinal Pseudo-obstruction

Bonora et al., 2015

ZDB-PUB-150113-12
PMID:25575569