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HGF Patterning Is Abnormal in Fog2 Null Mice In situ hybridization of HGF in E12.5 wild-type (A) and Fog2−/− (B) embryos demonstrates decreased expression in the region where the PPF meets the membranous diaphragm. Li, liver; Lu, lung.
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Synopsis
Birth defects involving the diaphragm are as common and as serious as genetic disorders such as cystic fibrosis, yet the underlying causes of these defects are unknown. Most babies born with diaphragmatic defects have very small lungs, and many die in the newborn period with severe breathing difficulties. It is unknown whether the small l...
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Purpose:
Pulmonary hypoplasia (PH), characterized by smaller lung size and reduced airway branching, remains a major therapeutic challenge in newborns with congenital diaphragmatic hernia (CDH). T-box transcription factors (Tbx) have been identified as key components of the gene network that regulates fetal lung development. Tbx2, Tbx4 and Tbx5 ar...
Citations
... The heart was the most enriched tissue, along with the lung, consistent with previous studies implicating FOG2 in lung development. 98 We assessed FOG2 genomic localization with respect to other known cardiac TFs. ChIP-seq suggested that FOG2 interacts with GATA TFs and other members of the mature cardiomyocyte TF kernel, including T-box TFs. ...
BACKGROUND
The relationship between heart failure (HF) and atrial fibrillation (AF) is clear, with up to half of patients with HF progressing to AF. The pathophysiological basis of AF in the context of HF is presumed to result from atrial remodeling. Upregulation of the transcription factor FOG2 (friend of GATA2; encoded by ZFPM2 ) is observed in human ventricles during HF and causes HF in mice.
METHODS
FOG2 expression was assessed in human atria. The effect of adult-specific FOG2 overexpression in the mouse heart was evaluated by whole animal electrophysiology, in vivo organ electrophysiology, cellular electrophysiology, calcium flux, mouse genetic interactions, gene expression, and genomic function, including a novel approach for defining functional transcription factor interactions based on overlapping effects on enhancer noncoding transcription.
RESULTS
FOG2 is significantly upregulated in the human atria during HF. Adult cardiomyocyte-specific FOG2 overexpression in mice caused primary spontaneous AF before the development of HF or atrial remodeling. FOG2 overexpression generated arrhythmia substrate and trigger in cardiomyocytes, including calcium cycling defects. We found that FOG2 repressed atrial gene expression promoted by TBX5 . FOG2 bound a subset of GATA4 and TBX5 co-bound genomic locations, defining a shared atrial gene regulatory network. FOG2 repressed TBX5-dependent transcription from a subset of co-bound enhancers, including a conserved enhancer at the Atp2a2 locus. Atrial rhythm abnormalities in mice caused by Tbx5 haploinsufficiency were rescued by Fog2 haploinsufficiency.
CONCLUSIONS
Transcriptional changes in the atria observed in human HF directly antagonize the atrial rhythm gene regulatory network, providing a genomic link between HF and AF risk independent of atrial remodeling.
... 97 Mice with a mutation in Fog2 present diaphragm abnormalities characterized by incomplete muscularization of the posterolateral diaphragm in 100% of mutant mice. 98 Regarding the Retinoid Hypothesis, both Gata4 and Fog2 are thought to be RA target genes, 84,87,99,100 and it has been shown that retinoids can regulate downstream gene expression by interacting with GATA4 and FOG2. 101 Interestingly, FOG2 has also been shown to interact with COUP-TF2, suggesting complex interactions with multiple transcription factors that are important in diaphragm development. ...
... 131 Similarly, FOG2 (OMIM: 603693) was originally linked to CDH as it is in the critical region 8q22-8q23, 127 with later studies showing specific mutations/ deletions of this gene were recurrently associated with CDH. 98,132 WT1, COUP-TF2, GATA4, and FOG2 are all expressed in the developing diaphragm, and all are strongly linked to CDH via both mouse and human mutations. All four genes have also been linked to RA signaling, providing supporting evidence for the importance of this pathway in normal diaphragm development. ...
Congenital diaphragmatic hernia (CDH) is a severe birth defect and a major cause of neonatal respiratory distress. Impacting ~2–3 in 10,000 births, CDH is associated with a high mortality rate, and long-term morbidity in survivors. Despite the significant impact of CDH, its etiology remains incompletely understood. In 2003, Greer et al. proposed the Retinoid Hypothesis, stating that the underlying cause of abnormal diaphragm development in CDH was related to altered retinoid signaling. In this review, we provide a comprehensive update to the Retinoid Hypothesis, discussing work published in support of this hypothesis from the past 20 years. This includes reviewing teratogenic and genetic models of CDH, lessons from the human genetics of CDH and epidemiological studies, as well as current gaps in the literature and important areas for future research. The Retinoid Hypothesis is one of the leading hypotheses to explain the etiology of CDH, as we continue to better understand the role of retinoid signaling in diaphragm development, we hope that this information can be used to improve CDH outcomes.
Impact
This review provides a comprehensive update on the Retinoid Hypothesis, which links abnormal retinoic acid signaling to the etiology of congenital diaphragmatic hernia.
The Retinoid Hypothesis was formulated in 2003. Twenty years later, we extensively review the literature in support of this hypothesis from both animal models and humans.
... Only a few of them seem likely to be associated with the development of the diaphragm based on our current understanding of molecular networks and signaling pathways throughout organogenesis. For example, molecules acting in retinol-pathways [7][8][9][10] or members of GATA4/FOG2-signaling which seem to be crucial for the non-muscular part of the premature diaphragm to evolve can relatively coherently be identified as definitive contributors to CDH [11][12][13][14][15]. Most of the described genetic anomalies, however, lack immediate association to diaphragm development; hence, their potential contribution to its pathogenicity can only be assumed with varying plausibility. ...
Congenital diaphragmatic hernia (CDH) is a major birth anomaly that often occurs with additional non-hernia-related malformations, and is then referred to as CDH+. While the impact of genetic alterations does not play a major role in isolated CDH, patients with CDH+ display mutations that are usually determined via array-based comparative genomic hybridization (aCGH). We analyzed 43 patients with CDH+ between 2012 and 2021 to identify novel specific mutations via aCGH associated with CDH+ and its outcome. Deletions (n = 32) and duplications (n = 29) classified as either pathological or variants of unknown significance (VUS) could be detected. We determined a heterozygous deletion of approximately 3.75 Mb located at 8p23.1 involving several genes including GATA4, NEIL2, SOX7, and MSRA, which was consequently evaluated as pathological. Another heterozygous deletion within the region of 9p23 (9,972,017-10,034,230 kb) encompassing the Protein Tyrosine Phosphatase Receptor Type Delta gene (PTPRD) was identified in 2 patients. This work expands the knowledge of genetic alterations associated with CDH+ and proposes two novel candidate genes discovered via aCGH.
... 1,2 Four classifications of CDH have been previously described (A,B,C,D) based on the size of the diaphragmatic defect and the co-occurrence of a cardiac defect (þ). 3 A variety of chromosomal and genomic alterations and sequence variants have been shown to cause or predispose to the development of CDH and may also have a direct effect on lung development. [4][5][6][7] Despite these discoveries, the genetic factors underling most cases of CDH remain unidentified. ...
FOXP1 encodes a transcription factor involved in tissue regulation and cell-type-specific functions. Haploinsufficiency of FOXP1 is associated with a neurodevelopmental disorder: autosomal dominant mental retardation with language impairment with or without autistic features. More recently, heterozygous FOXP1 variants have also been shown to cause a variety of structural birth defects including central nervous system (CNS) anomalies, congenital heart defects, congenital anomalies of the kidney and urinary tract, cryptorchidism, and hypospadias. In this report, we present a previously unpublished case of an individual with congenital diaphragmatic hernia (CDH) who carries an approximately 3.8 Mb deletion. Based on this deletion, and deletions previously reported in two other individuals with CDH, we define a CDH critical region on chromosome 3p13 that includes FOXP1 and four other protein-coding genes. We also provide detailed clinical descriptions of two previously reported individuals with CDH who carry de novo, pathogenic variants in FOXP1 that are predicted to trigger nonsense-mediated mRNA decay. A subset of individuals with putatively deleterious FOXP4 variants has also been shown to develop CDH. Since FOXP proteins function as homo- or heterodimers and the homologs of FOXP1 and FOXP4 are expressed at the same time points in the embryonic mouse diaphragm, they may function together as a dimer, or in parallel as homodimers, to regulate gene expression during diaphragm development. Not all individuals with heterozygous, loss-of-function changes in FOXP1 develop CDH. Hence, we conclude that FOXP1 acts as a susceptibility factor that contributes to the development of CDH in conjunction with other genetic, epigenetic, environmental, and/or stochastic factors.
... Defects in the development of the PPFs will lead to incomplete development Perveen et al. 10.3389/fped.2022.932463 of the diaphragm. Several genes associated with CDH are expressed in relation to PPFs and may be the cause of incomplete diaphragmatic development (17,18). The genetic inactivation of GATA-4 in mice showed the development of localized regions that lack muscle progenitors leading to amuscular patches in the developing diaphragm that are mechanically weaker resulting in herniation. ...
... This interaction modulates the transcriptional activity of GATA4 (Bashamboo et al., 2014;De Luca et al., 2011). Heterozygous variants in ZFPM2 are associated with conotruncal heart defects, specifically tetralogy of Fallot, double outlet right ventricle, and transposition of the great arteries, as well as a congenital diaphragmatic hernia (Ackerman et al., 2005;De Luca et al., 2011). Heterozygous variants in the ZFPM2 gene have also been reported in individuals with 46,XY disorders of sex development (Bashamboo et al., 2014) like with GATA4 (Martinez de LaPiscina et al., 2018). ...
Aldehyde Dehydrogenase 1, Family Member A2 (ALDH1A2) is essential for the synthesis of retinoic acid from vitamin A. Studies in model organisms demonstrate a critical role for ALDH1A2 in embryonic development, yet few pathogenic variants are linked to congenital anomalies in humans. We present three siblings with multiple congenital anomaly syndrome linked to biallelic sequence variants in ALDH1A2. The major congenital malformations affecting these children include tetralogy of Fallot, absent thymus, diaphragmatic eventration, and talipes equinovarus. Upper airway anomalies, hypocalcemia, and dysmorphic features are newly reported in this manuscript. In vitro functional validation of variants indicated that substitutions reduced the expression of the enzyme. Our clinical and functional data adds to a recent report of biallelic ALDH1A2 pathogenic variants in two families with a similar constellation of congenital malformations. These findings provide further evidence for an autosomal recessive ALDH1A2-deficient recognizable malformation syndrome involving the diaphragm, cardiac and musculoskeletal systems.
... For example, a trio exome/ WES analysis revealed that the GATA4, ZFPM2, and GATA6 genes are associated with CDH (Yu et al. 2013, Longoni et al. 2014Yu et al. 2020). In particular, GATA6, ZFMP2, GATA4, SYNC, NR2F2, EYA1, CTNNB1, and FGfrL1 genes have been demonstrated to show considerable changes in expression in CDH patients (Ameis et al. 2017, Yu et al. 2013, Longoni et al. 2014, Ackerman et al. 2005, Kammoun et al. 2018, You et al. 2005, Paris et al. 2015, Amann et al. 2014. ...
... Herein, we found that expression of GATA6 considerably upregulated in two CDH patients. Ackerman et al. reported that CDH and bilateral pulmonary hypoplasia occurred as a result of mutations in the FOG2 (ZFPM2) gene resulting in N-etHyl-N-nitrosourea (Ackerman et al. 2005). We also indicated that FOG2 (ZFPM2) expression is significantly increased in all patients. ...
Congenital diaphragmatic hernia (CDH) is an anomaly characterized by a defect in the diaphragm, leading to the passage of intra-abdominal organs into the thoracic cavity. Herein, the presented work analyzes the global gene expression profiles in nine CDH and one healthy newborn. All of the patients had left posterolateral (Bochdalek) diaphragmatic hernia, operated via an abdominal approach, and stomach and bowels in the thorax cavity. Some patients also had additional anomalies. A total of 560 differentially regulated genes were measured. Among them, 11 genes showed significant changes in expression associated with lung tissue, vascular structure development, and vitamin A metabolism, which are typical ontologies related to CDH etiology. Among them, SLC25A24 and RAB3IL1 are involved in angiogenesis, HIF1A and FOXC2-AS1 are related with the alveolus, MAGI2-AS3 is associated with the diaphragm, LHX4 and DHH are linked with the lung, and BRINP1, FZD9, WNT4, and BLOC1S1-RDH5 are involved in retinol. Besides, the expression levels of some previously claimed genes with CDH etiology also showed diverse expression patterns in different patients. All these indicated that CDH is a complex, multigenic anomaly, requiring holistic approaches for its elucidation.
... Zinc finger protein 2 (ZFPM2), formerly known as friend of GATA-binding protein 2 (FOG2), encodes another zinc fingercontaining protein that regulates the transcriptional activity of GATA4, hereby controlling a number of developmental mechanisms in the forming diaphragm and lung (78)(79)(80)(81). In mice, Fog2 was initially found to be expressed in the embryonic septum transversum of the diaphragm (81). ...
Congenital diaphragmatic hernia (CDH) is a relatively common and life-threatening birth defect, characterized by an abnormal opening in the primordial diaphragm that interferes with normal lung development. As a result, CDH is accompanied by immature and hypoplastic lungs, being the leading cause of morbidity and mortality in patients with this condition. In recent decades, various animal models have contributed novel insights into the pathogenic mechanisms underlying CDH and associated pulmonary hypoplasia. In particular, the generation of genetically modified mouse models, which show both diaphragm and lung abnormalities, has resulted in the discovery of multiple genes and signaling pathways involved in the pathogenesis of CDH. This article aims to offer an up-to-date overview on CDH-implicated transcription factors, molecules regulating cell migration and signal transduction as well as components contributing to the formation of extracellular matrix, whilst also discussing the significance of these genetic models for studying altered lung development with regard to the human situation.
... Mutations in many GATA family members have resulted in congenital heart defects, including atrial septal defects, and variants in some GATA genes can also cause CDH (Lin et al., 2010;Maitra et al., 2010;Tomita-Mitchell et al., 2007;Wang et al., 2011;Yu et al., 2013Yu et al., , 2014Zhang et al., 2008). Heterozygous loss-of-function variants in ZFPM2 have been previously associated with CDH and cardiac malformations (Ackerman et al., 2005;Bleyl et al., 2007;Luca et al., 2011;Tan et al., 2012), establishing haploinsufficiency as the molecular mechanism for pathogenesis. Given the known association between CDH and ZFPM2, we conclude that this complex variant explains the CDH and cardiac phenotypes shared by the proband and mother. ...
Background:
Genetic disorders contribute to significant morbidity and mortality in critically ill newborns. Despite advances in genome sequencing technologies, a majority of neonatal cases remain unsolved. Complex structural variants (SVs) often elude conventional genome sequencing variant calling pipelines and will explain a portion of these unsolved cases.
Methods:
As part of the Utah NeoSeq project, we used a research-based, rapid whole-genome sequencing (WGS) protocol to investigate the genomic etiology for a newborn with a left-sided congenital diaphragmatic hernia (CDH) and cardiac malformations, whose mother also had a history of CDH and atrial septal defect.
Results:
Using both a novel, alignment-free and traditional alignment-based variant callers, we identified a maternally inherited complex SV on chromosome 8, consisting of an inversion flanked by deletions. This complex inversion, further confirmed using orthogonal molecular techniques, disrupts the ZFPM2 gene, which is associated with both CDH and various congenital heart defects.
Conclusions:
Our results demonstrate that complex structural events, which often are unidentifiable or not reported by clinically validated testing procedures, can be discovered and accurately characterized with conventional, short-read sequencing and underscore the utility of WGS as a first-line diagnostic tool.
... Defects in the development of the PPFs are a significant source of CDH. Most CDH-implicated genes that have been examined are expressed in the PPF cells (7,(12)(13)(14)(15). Mutations in CDH-implicated genes can lead to the incomplete expansion of the PPFs and thus lead to incompletely developed diaphragms that allow herniation of abdominal contents into thoracic cavity [e.g., (16)]. ...
Congenital diaphragmatic hernia (CDH) is a structural birth defect characterized by a diaphragmatic defect, lung hypoplasia and structural vascular defects. In spite of recent developments, the pathogenesis of CDH is still poorly understood. CDH is a complex congenital disorder with multifactorial etiology consisting of genetic, cellular and mechanical factors. This review explores the cellular origin of CDH pathogenesis in the diaphragm and lungs and describes recent developments in basic and translational CDH research.
