Refining critical regions in 15q24 microdeletion syndrome pertaining to autism

1 INTRODUCTION

Chromosome 15q24 microdeletion syndrome is a rare syndrome (Sharp et al., 2007), characterized by distinct phenotypes of global developmental delay, intellectual disability, facial dysmorphisms, congenital malformations of the hands and feet, eye, ear and genitalia, and joint laxity. The phenotypes are highly variable across different deletion carriers with >75% patients presenting facial features, developmental delay/intellectual disability, significant speech delay and hypotonia, 50–75% patients manifesting eye abnormalities, most frequently strabismus, digital anomalies, ear anomalies; 25–50% showing joint laxity, hearing loss, abnormalities on brain magnetic resonance imaging (MRI), hypospadias, and/or micropenis; 10–25% having seizures, sensorineural hearing loss, and conductive hearing loss; <10% featuring heart defects, diaphragmatic hernia, bowel atresia, and Pierre Robin syndrome (Mefford et al., 2012a, 2012b). It was first defined in 2007, with many more cases now reported and the prevalence assessed to be 3:10,000–4:10,000 ([Klopocki et al., 2008; Marshall et al., 2008; Kiholm Lund, et al., 2008; Andrieux et al., 2009; El-hattab et al., 2009; Masurel-Paulet et al., 2009, Van Esch et al., 2009; El-Hattab et al., 2010; McInnes et al., 2010; Ng et al., 2011; Brun et al., 2012; Mefford et al., 2012a, 2012b; Narumi et al., 2012; Gao et al., 2014; Samuelsson et al., 2015; Siu etal.,2016; Palazon-Bru et al., 2016; Romano et al., 2017; Ahram et al., 2017; Huynh et al.,2017; Dai et al., 2017; Wang et al., 2019]). Most of 15q24 deletions were found between 1.7 and 6.1 Mb in size with breakpoints localized to the clusters of five low copy regions (LCRs) from centromere to telomere, which were identified and designated as LCR15q24A (BP4), B (BP1), C, D (BP2), and E (BP3). Out of the cases, a few manifesting an additional phenotype of autism spectrum disorder (ASD; Ahram et al., 2017; McInnes et al., 2010; Mefford et al., 2012a, 2012b; Siu et al., 2016) carried mostly atypical deletions at 15q24 lying outside the critical region (Huynh et al., 2018; McInnes et al., 2010).

ASD is a vastly heterogeneous group of highly heritable neurodevelopmental disorders, characterized by impairments in reciprocal social interaction and communication, restricted and repetitive behaviors and interests, affecting ~1% of the population with the age of onset usually before 3 years (Carter & Scherer, 2013; Hoang, Cytrynbaum, & Scherer, 2018; Zhang et al., 2018). Various genetic mutations have been known to be implicated in ASD, ranging from chromosome abnormalities and copy-number variations (CNVs) to single-nucleotide variations (SNVs; Liu et al., 2015; Mak et al., 2017; Yuen et al., 2017). In 2007, the Autism Genome Project Consortium proposed the location of 15q23-25.3 would be the autism susceptibility locus by linkage and CNV analyses of 1,181 families with at least two ASD patients (Autism Genome Project Consortium, et al., 2007). Since then, more autistic patients have been reported carrying the 15q24 deletions (Ahram et al., 2017; Marshall et al., 2008; McInnes et al., 2010; Mefford et al., 2012a, 2012b; Siu et al., 2016). Here, we reported a de novo 2.58 Mb deletion at 15q24.1-q24.2 in a boy with ASD, and other phenotypes overlapped with those previously reported cases of 15q24 microdeletion syndrome. This is one case of a family-based study with the purpose of identifying genetic variants related to ASD. We reviewed all the available cases with 15q24 CNVs, focusing on microdeletions located at 15q24 in the published literature, DECIPHER (Firth et al., 2009) and ClinGen (Rehm et al., 2015; Solomon, Nguyen, Bear, & Wolfsberg, 2013) databases. We observed a second critical region of 650-kb interval between LCR15q24A and 15q24B that might contribute genetically to the ASD often found in this syndrome.

2 MATERIALS AND METHODS

The Review Board of Qilu Children's Hospital of Shandong University has approved this study. The patient's parents consented to this study.

3 RESULTS

3.3 Identifying the critical region

To define the minimal critical region and genes that might contribute to the syndrome, we reviewed all the cases with isolated 15q24 deletions and duplications. In total, 46 deletions and 12 duplications were found including three embryos of miscarriage from literature, and an additional 16 deletions were from DECIPHER database, which were summarized in Tables S1–S3, separately. The clinical features were compared and the occurrence of each phenotype was calculated for all cases with 15q24 deletions except that case of embryo (Table 1). The CNV map of all cases was represented graphically in Figure 1 with two highlighted minimal critical regions. The first critical region is the one previously defined (chr15:74,500,000–75,700,000; 15q24B-D; first critical region) of a 1.2 Mb interval involving 52 (83.9%) cases of deletions and six (50.0%) cases of duplications. This region is gene-rich encompassing 39 genes including some OMIM genes (e.g., STRA6, CYP11A1, SEMA7A, ARID3B, CPLK3, CSK, and SIN3A; Magoulas & El-Hattab, 2012). This locus is more likely associated with other phenotypes of the syndrome excluding ASD. The second critical region is predicted from chr15: 73,010,000–73,660,000 (second critical region) covering 650-kb implicating 37 (59.7%) cases of deletions and three (25%) cases of duplications, including four genes (BBS4, ADPGK, NEO1, and HCN4), which presumably contributed to the phenotype of ASD of this syndrome.

4 DISCUSSION

In this study, we proposed the second critical region of a 650-kb interval located between 15q24A and 15q24B based on cases study and review, which might contribute to the ASD phenotype of 15q24 microdeletion syndrome, while the first critical region of a 1.2-Mb span located between 15q24B and 15q24D defined previously is more likely associated with the other phenotypes of the syndrome, such as developmental delay, facial dysmorphism, recurrent infection, and congenital malformations (Magoulas & El-Hattab, 2012). Our case carried a de novo deletion of 2.58-Mb located between 15q24A and 15q24C presented the common phenotypes, such as severe speech problem, congenital anomalies, and additional ASD which is mostly attributed by the second critical region and the potential candidate gene of NEO1.

The smallest region of overlap has been predicted to be an approximately 1.2-Mb region between 15q24B and 15q24D as a majority of 19 patients with 15q24 deletions involved in this region (Magoulas & El-Hattab, 2012; McInnes et al., 2010; Mefford et al., 2012a, 2012b). So far, more cases with 15q24 deletions and duplications have been reported to present common clinical features (Roggenbuck et al., 2004; Miller et al., 2005; Schluth et al., 2005; Andrieux et al., 2009; El-Hattab et al., 2010; Roetzer et al., 2010; Cukier et al., 2011; Kim et al., 2011; O'Connor et al., 2011; Huynh et al., 2018; Ochando et al., 2018; Mefford et al., 2012b; Siu et al., 2016). To increase the number of cases, we searched all the literature, DECIPHER, and ClinGen databases for 15q24 deletions and duplications. By comparing the locus of CNVs linked to the clinical features, an interesting finding was noticed that the 25 cases carrying deletions located between 15q24B and 15q24E only covering the first critical region and excluding 15q24A and 15q24B did not present ASD phenotypes. Furthermore, all the ASD cases including our new patient in this study involved the deletions between 15q24A and 15q24B. Thus, we defined the minimal region within it as the second critical region, which is 650-kb in size including four genes, that is, BBS4, ADPGK, NEO1, and HCN4.

Previous studies have shown that homozygous loss-of-function mutations in some genes within first critical region of chr15q24 (see Figure 1) are associated with different congenital defects, such as STRA6 associated with Matthew-Wood syndrome presenting pulmonary agenesis/hypoplasia, congenital cardiac defect, microphthalmia/anophthalmia, diaphragmatic hernias, and urogenital malformations (Sadowski et al., 2017); ARID3B has been reported as a candidate gene associated with congenital cardiac malformations (Uribe et al., 2014); CYP11A, a cholesterol side chain cleavage enzyme associated with congenital lipoid adrenal hyperplasia (Horikawa, 2004); MPI related to congenital disorder of glycation and hyperinsulinemic hypoglycemia (Deeb & Al Amoodi, 2018); EDC3 related to autosomal recessive intellectual disability (Ahmed et al., 2015), indicating the haploinsufficiency of these genes might contribute to the corresponding phenotypes of the syndrome.

It has been known that some genes in first critical region of chr15q24 associated with the development of nervous system, such as SIN3A, SEMA7A, and CPLX3 (Cioli, Abdi, Beaton, Burnod, & Mesmoudi, 2014; Pasterkamp, Peschon, Spiggs, & Kolodkin, 2003; Witteveen et al., 2016), which suggests there might be potentially synergistic effects in these genes underlie the psychiatric phenotypes. In addition, two genes of NEIL1 and CSK associated with immunodeficiency, and T cell antigen receptor signaling (Lee et al., 2016; Manz et al., 2015; Romano et al., 2017; Sun et al., 2018; Zhang, Ren, Yang, & Ding, 2016).

By analyzing the clinical phenotypes of the cases implicated in 15q24, we defined the second critical region covering 650 kb located between 15q24A and 15q24B, encompassing four genes of BBS4, NEO1, HCN4, and ADPGK, which have been described to be associated with the atypical 15q24 deletion syndrome (Huynh et al., 2018; McInnes et al., 2010). BBS4 encoding Bardet-Biedl syndrome (BBS) 4 protein, is the causative gene of BBS that is an autosomal recessive ciliopathy characterized by obesity, polydactyly, renal malformation, rod-cone dystrophy, genital abnormalities, and cognitive disability (Ece Solmaz et al., 2015). Silencing of BBS4 promotes cell division and aberrant differentiation which results in adipose tissue formation underlying the BBS obesity (Aksanov, Green, & Birk, 2014).

NEO1, a membrane receptor, regulates pivotal developmental processes in the central nervous system of embryos involving interneuron migration, axon guidance, and cell death, which have been proved to contribute the autism pathophysiology (Fitzgerald, Cole, Hammond, Seaman, & Cooper, 2006; Matsunaga et al., 2004; Mawdsley et al., 2004; Wilson & Key, 2006). Siu et al. identified genetic changes in NEO1 gene from 2 out of 66 ASD patients and found the NEO1 played a critical role in cortical interneuron development by function experiment (Siu et al., 2016). Therefore, we proposed it might be a novel ASD-associated gene. HCN4 is a hyperpolarization-activated cyclic nucleotide gated channel 4 expressed predominantly in heart ventricle, atrium, and neurons and has been reported being associated with Sick Sinus Syndrome and cardiac abnormalities (Ishikawa et al., 2017; Milano et al., 2014). Recent studies showed it is highly expressed in habenula and thalamus, but is low expression level in amygdala, cortex, and hippocampus, which are known ASD associated regions. (Oyrer et al., 2019; Seo et al., 2015) and a loss of function mutation in HCN4 is associated with development of infantile epilepsy (Campostrini et al., 2018). ADPGK is ADP-dependent glucokinase and an alternative, glycolytic enzyme mediating generation of the oxidative signal whose downregulation inhibits oxidative signal and induces NF-kB-dependent gene expression, which makes T cell activation (Kaminski et al., 2012). We summarized the evidence of genes in the second region as Table 3. After studied, the phenotypes of the patients involved in this region, we predicted that the second critical region might be mainly responsible for behavior problems like ASD, but more studies are needed for the exact function of the potential critical genes within this region.

This study presents additional evidence of 15q24 microdeletion syndrome with clinical and molecular findings in a patient with ASD. Thorough review of the cases and this new patient suggests that the second critical region of 15q24 and genes, particularly NEO1, play an etiologic role on the ASD phenotype of the syndrome.

CONFLICT OF INTEREST

S.W.S. is on the Scientific Advisory Committee of Population Diagnostics and Deep Genomics, and Lineagen Corp. Licenses intellectual property relevant to autism testing held by the Hospital for Sick Children.

AUTHOR CONTRIBUTIONS

Y.L., S.W.S., and Z.G. conceived and designed the study. X.Y. and Y.L. conducted the experiments. Y.L., M.Z., and S.W.S. analyzed the data. Y.Z., R.D., D.Z., and Z.G. contributed clinical diagnosis of the patients. Y.L. and M.Z. wrote the manuscript.