Genetic underpinnings explored: OPA1 deletion and complex phenotypes on chromosome 3q29.

BACKGROUND: Copy number variations (CNVs) have emerged as significant contributors to the elusive genetic causality of inherited eye diseases. In this study, we describe a case with optic atrophy and a brain aneurysm, in which a de novo CNV 3q29 deletion was identified. CASE PRESENTATION: A 40-year-old female patient was referred to our department after undergoing aneurysm transcatheter arterial embolization for a brain aneurysm. She had no history of systemic diseases, except for unsatisfactory best-corrected visual acuity (BCVA) since elementary school. Electrophysiological tests confirmed the findings in retinal images, indicating optic nerve atrophy. Chromosomal microarray analysis revealed a de novo deletion spanning 960 kb on chromosome 3q29, encompassing OPA1 and six neighboring genes. Unlike previously reported deletions in this region associated with optic atrophy, neuropsychiatric disorders, and obesity, this patient displayed a unique combination of optic atrophy and a brain aneurysm. However, there is no causal relationship between the brain aneurysm and the CNV. CONCLUSION: In conclusion, the optic atrophy is conclusively attributed to the OPA1 deletion, and the aneurysm could be a coincidental association. The report emphasizes the likelihood of underestimating OPA1 deletions due to sequencing technology limitations. Recognizing these constraints, healthcare professionals must acknowledge these limitations and consistently search for OPA1 variants/deletions in Autosomal Dominant Optic Atrophy (ADOA) patients with negative sequencing results. This strategic approach ensures a more comprehensive exploration of copy-number variations, ultimately enhancing diagnostic precision in the field of genetic disorders.

Clinical Characteristics and Genetic Variants in Taiwanese Patients With PROM1-Related Inherited Retinal Disorders.

Purpose: This study investigated the clinical characteristics of patients with PROM1-related inherited retinal diseases (IRDs). Methods: Patients diagnosed with IRDs who had mutations in PROM1 were identified at Linkou Chang Gung Memorial Hospital and Kaohsiung Medical University Hospital in Taiwan. Information on clinical characteristics and best-corrected visual acuity was recorded. Color fundus (CF) images, fundus autofluorescence photography (FAF), spectral-domain optical coherence tomography (SD-OCT), and electroretinograms (ERGs) were analyzed to examine patient phenotypes. PROM1 variants were detected using whole exome sequencing and verified by Sanger sequencing. Results: Fourteen patients from nine families with PROM1-related IRDs were analyzed. Most patients exhibited chorioretinal atrophy in the macular area, with or without extramacular involvement on CF. Similarly, hypo-autofluorescence confined to the macular area, with or without extramacular involvement, was present for most patients on FAF. Furthermore, SD-OCT revealed outer retinal tubulations and focal or diffuse retinal thinning. ERGs showed variable findings, including maculopathy with normal ERG, subnormal cone response, and extinguished rod and cone responses. We detected five variants of the PROM1 gene, including c.139del, c.794del, c.1238T>A, c.2110C>T, and c.1117C>T. Conclusions: In this study, we evaluated 14 Taiwanese patients with five PROM1 variants. Additionally, incomplete penetrance of heterozygous PROM1 variants was observed. Furthermore, patients with autosomal dominant PROM1 variants had lesions in the macular area and the peripheral region of the retina. SD-OCT serves as a useful tool for early detection of PROM1-related IRDs, as it captures certain signs of such diseases.

Maternal mosaicism in SSBP1 causing optic atrophy with retinal degeneration: implications for genetic counseling.

BACKGROUND: Optic atrophy-13 with retinal and foveal abnormalities (OPA13) (MIM #165510) is a mitochondrial disease in which apparent bilateral optic atrophy is present and sometimes followed by retinal pigmentary changes or photoreceptors degeneration. OPA13 is caused by heterozygous mutation in the SSBP1 gene, associated with variable mitochondrial dysfunctions. RESULTS: We have previously reported a 16-year-old Taiwanese male diagnosed with OPA13 and SSBP1 variant c.320G>A (p.Arg107Gln) was identified by whole exon sequence (WES). This variant was assumed to be de novo since his parents were clinically unaffected. However, WES and Sanger sequencing further revealed the proband's unaffected mother carrying the same SSBP1 variant with a 13% variant allele frequency (VAF) in her peripheral blood. That finding strongly indicates the maternal gonosomal mosaicism contributing to OPA13, which has not been reported before. CONCLUSIONS: In summary, we described the first case of OPA13 caused by maternal gonosomal mosaicism in SSBP1. Parental mosaicism could be a serious issue in OPA13 diagnosis, and appropriate genetic counseling should be considered.

Maternal Mosaicism in SSBP1 Causing Optic Atrophy with Retinal Degeneration: Implications for Genetic Counseling.

Background: Optic atrophy-13 with retinal and foveal abnormalities (OPA13) (MIM #165510) is a mitochondrial disease in which apparent bilateral optic atrophy is present and sometimes followed by retinal pigmentary changes or photoreceptors degeneration. OPA13 is caused by heterozygous mutation in the SSBP1 gene, associated with variable mitochondrial dysfunctions. Results: We have previously reported a 16-year-old Taiwanese male diagnosed with OPA13 and SSBP1 variant c.320G>A (p.Arg107Gln) was identified by whole exon sequence (WES). This variant was assumed to be de novo since his parents were clinically unaffected. However, WES and Sanger sequencing further revealed the proband’s unaffected mother carrying the same SSBP1 variant with a 13% variant allele frequency (VAF) in her peripheral blood. That finding strongly indicates the maternal gonosomal mosaicism contributing to OPA13, which has not been reported before. Conclusions: In summary, we described the first case of OPA13 caused by maternal gonosomal mosaicism in SSBP1 . Parental mosaicism could be a serious issue in OPA13 diagnosis, and appropriate genetic counseling should be considered.

Investigation into the genetics of fetal congenital lymphatic anomalies.

OBJECTIVE: Congenital lymphatic anomalies (LAs) arise due to defects in lymphatic development and often present in utero as pleural effusion, chylothorax, nuchal and soft tissue edema, ascites, or hydrops. Many LAs are caused by single nucleotide variants, which are not detected on routine prenatal testing. METHODS: Demographic data were compared between two subcohorts, those with clinically significant fetal edema (CSFE) and isolated fetal edema. A targeted variant analysis of LA genes was performed using American College of Medical Genetics criteria on whole exome sequencing (WES) data generated for 71 fetal edema cases who remained undiagnosed after standard workup. RESULTS: CSFE cases had poor outcomes, including preterm delivery, demise, and maternal preeclampsia. Pathogenic and likely pathogenic variants were identified in 7% (5/71) of cases, including variants in RASopathy genes, RASA1, SOS1, PTPN11, and a novel PIEZO1 variant. Variants of uncertain significance (VOUS) were identified in 45% (32/71) of cases. In CSFEs, VOUS were found in CELSR1, EPHB4, TIE1, PIEZO1, ITGA9, RASopathy genes, SOS1, SOS2, and RAF1. CONCLUSIONS: WES identified pathogenic and likely pathogenic variants and VOUS in LA genes in 51% of fetal edema cases, supporting WES and expanded hydrops panels in cases of idiopathic fetal hydrops and fluid collections.

Congenital Stationary Night Blindness: Clinical and Genetic Features.

Congenital stationary night blindness (CSNB) is an inherited retinal disease (IRD) that causes night blindness in childhood with heterogeneous genetic, electrophysical, and clinical characteristics. The development of sequencing technologies and gene therapy have increased the ease and urgency of diagnosing IRDs. This study describes seven Taiwanese patients from six unrelated families examined at a tertiary referral center, diagnosed with CSNB, and confirmed by genetic testing. Complete ophthalmic exams included best corrected visual acuity, retinal imaging, and an electroretinogram. The effects of identified novel variants were predicted using clinical details, protein prediction tools, and conservation scores. One patient had an autosomal dominant CSNB with a RHO variant; five patients had complete CSNB with variants in GRM6, TRPM1, and NYX; and one patient had incomplete CSNB with variants in CACNA1F. The patients had Riggs and Schubert-Bornschein types of CSNB with autosomal dominant, autosomal recessive, and X-linked inheritance patterns. This is the first report of CSNB patients in Taiwan with confirmed genetic testing, providing novel perspectives on molecular etiology and genotype-phenotype correlation of CSNB. Particularly, variants in TRPM1, NYX, and CACNA1F in our patient cohort have not previously been described, although their clinical significance needs further study. Additional study is needed for the genotype-phenotype correlation of different mutations causing CSNB. In addition to genetic etiology, the future of gene therapy for CSNB patients is reviewed and discussed.

Case report: novel PCDH15 variant causes usher syndrome type 1F with congenital hearing loss and syndromic retinitis pigmentosa.

BACKGROUND: Usher syndrome (USH) is an autosomal recessive disorder primarily responsible for deaf-blindness. Patients with subtype Usher syndrome type 1 (USH1) typically experience congenital sensorineural hearing loss, abnormal vestibular function, and retinitis pigmentosa (RP). Here we present a case of Usher syndrome type 1F (USH1F) with a novel homozygous variant in the calcium-dependent cell-cell adhesion protocadherin-15 (PCDH15) gene. CASE PRESENTATION: Ophthalmic examinations were evaluated over a course of 10 years and the disease-causing variant was identified by whole exome sequencing (WES). Initial and follow-up examination of color fundus photos after 10 years revealed an increase in bone spicule pigment deposits in both eyes. A parafoveal hyper-AF ring in both eyes was shown in fundus autofluorescence (FAF) with a progressive diameter-wise constriction observed over 8 years. Outer nuclear layer (ONL) loss was observed in parafoveal and perifoveal regions of both eyes on spectral domain-optical coherence tomography (SD-OCT). Full-field electroretinography (ffERG) showed extinguished global retinal function. WES identified a novel two-base-pair deletion, c.60_61del (p.Phe21Ter), in the PCDH15 gene, confirming the diagnosis of USH1F. CONCLUSIONS: We report a novel homozygous PCDH15 pathogenic variant expected to lead to nonsense-mediated decay (NMD) of PCDH15 mRNA. The patient exhibits a loss of function with USH1F, experiencing congenital hearing loss and syndromic RP.

Glycated Hemoglobin < 6.5% Is Associated With Uroseptic Shock in Diabetic Patients With Urinary Tract Infection.

This study aimed to compare the clinical characteristics and treatment outcomes of diabetic and non-diabetic individuals with urinary tract infection (UTI) and determine whether glycated hemoglobin (HbA1c) levels <6. 5% leads to uroseptic shock in diabetic individuals. We retrospectively collected and analyzed the clinical data of 1,363 individuals with UTIs in Taiwan from January 2006 to January 2018. Of the 345 diabetic individuals, 61 (17.7%) developed uroseptic shock. Diabetic patients who developed uroseptic shock tended to be older and males and, had a history of congestive heart failure, urolithiasis, higher serum creatinine level during hospitalization, lower serum HbA1c level, bacteremia, and acute kidney injury. Backward stepwise multivariate logistic regression analysis showed that male gender [odds ratio (OR), 1.861; 95% confidence interval (CI), 1.009-3.433; P = 0.047], congestive heart failure (OR, 4.036; 95% CI, 1.542-10.565; P = 0.004), bacteremia (OR, 2.875; 95% CI, 1.539-5.370; P = 0.001), and HbA1c level <6.5% (OR, 2.923; 95% CI, 1.580-5.406; P = 0.001) were associated with an increased risk of developing uroseptic shock among diabetic patients during hospitalization due to UTI. HbA1c level <6.5% is independently associated with uroseptic shock in diabetic patients with UTI.

Risk factors for uroseptic shock in hospitalized patients aged over 80 years with urinary tract infection.

BACKGROUND: The purpose of this study was to compare clinical characteristics and outcomes in individuals of different age groups with urinary tract infection (UTI), and to identify the relationships among advanced age and uroseptic shock. METHODS: This retrospective study compared clinical characteristics and outcomes in patients in different age groups with UTI and identified relationships between advanced age and uroseptic shock among hospitalized patients with UTI in an acute hospital care setting from January 2006 to October 2018. Patients were divided into young (age below 65 years), old (65-80 years), and very old (above 80 years) groups. RESULTS: Of 1,043 participants, 269 (25.8%) were very old and 200 (19.2%) developed uroseptic shock. Very old age [odds ratio (OR) 1.99, 95% confidence interval (CI): 1.25-3.19, P=0.004], male (OR 1.54, 95% CI: 1.07-2.24, P=0.022), presented flank pain (OR 1.54, 95% CI: 1.05-2.24, P=0.025), congestive heart failure (CHF) (OR 2.54, 95% CI: 1.27-5.06, P=0.008), acute kidney injury (AKI) (OR 4.19, 95% CI: 2.78-6.30, P<0.001), bacteremia (OR 1.78, 95% CI: 1.25-2.53, P=0.001), and multiple drug-resistant (MDR) bacteria (OR 1.43, 95% CI: 1.02-2.00, P=0.039) were associated with an increased risk of uroseptic shock in patients with UTI. In very old patients with UTI, bacteremia (OR 2.54, 95% CI: 1.38-4.69, P=0.003) and AKI (OR 4.37, 95% CI: 2.15-8.90, P<0.001) were independently associated with uroseptic shock. CONCLUSIONS: Very old patients with UTI had a higher risk of developing uroseptic shock than younger patients. Bacteremia was an independent risk factor for uroseptic shock in very old patients with UTI.

Molecular profiling of gynecologic cancers for treatment and management of disease - demonstrating clinical significance using the AMP/ASCO/CAP guidelines for interpretation and reporting of somatic variants.

Molecular features of gynecologic cancers have been investigated in comprehensive studies, but correlation of these molecular signatures with clinical significance for precision medicine is yet to be established. Towards this end, we evaluated 95 gynecologic cancer cases submitted for testing using The JAX ActionSeq™ NGS panel. Molecular profiles were studied and compared to TCGA datasets to identify similarities and distinguishing features among subtypes. We identified 146 unique clinically significant variants (Tier I and II) across 45 of the 212 genes (21%), in 87% (83/95) of cases. TP53, PTEN, ARID1A, PIK3CA and ATM were the most commonly mutated genes; CCNE1 and ERBB2 amplifications were the most frequently detected copy-number alterations. PARP inhibitors were among the most commonly reported drug class with clinical trials, consistent with the frequency of DNA damage-response pathway mutations in our cohort. Overall, our study provides additional insight into the molecular profiles of gynecologic cancers, highlighting regulatory pathways involved, raising the potential implications for targeted therapeutic options currently available.

Molecular profiling of CNS tumors for the treatment and management of disease.

The World Health Organization (WHO) has defined more than 130 distinct central nervous system (CNS) tumor entities, of which glioblastoma is the most fatal primary brain tumor. However, the correlation of the molecular signatures of glioblastoma with clinical significance for precision medicine is not well-known. How, and to what extent these variants may affect clinical decision making remains uncertain. Here, we evaluate 48 glioblastomas submitted for testing using the JAX ActionSeq™ Next-generation sequencing (NGS) panel. We identified 131 clinically significant variants (Tier I and II) across 30 of the 212 genes (14%). TP53, EGFR, PTEN, IDH1 were the most commonly mutated genes; EGFR, CDK4 amplifications, and CDKN2A deletion were the most frequently detected copy-number alterations. CDK4/6 and PI3K inhibitors were among the most commonly reported drug class with FDA approved therapies and investigational therapies, which is consistent with the frequencies of these genes in our cohort. Overall, our study established the molecular profiles of glioblastoma based on the 2017 joint consensus guidelines by AMP/ASCO/CAP and provides the potential implications for targeted therapeutic options currently available.

Mutation Yield of a Custom 212-Gene Next-Generation Sequencing Panel for Solid Tumors: Clinical Experience of the First 260 Cases Tested Using the JAX ActionSeq™ Assay.

OBJECTIVE: The study aimed to retrospectively evaluate the positive yield rate of a custom 212-gene next-generation sequencing (NGS) panel, the JAX ActionSeq™ assay, used in molecular profiling of solid tumors for precision medicine. METHODS: We evaluated 261 cases tested over a 24-month period including cancers across 24 primary tissue types and report on the mutation yield in these cases. RESULTS: Thirty-three of the 261 cases (13%) had no detectable clinically significant variants. In the remaining 228 cases (87%), we identified 550 clinically significant variants in 88 of the 212 genes, with four of fewer clinically significant variants being detected in 62 of 88 genes (70%). TP53 had the highest number of variants (125), followed by APC (47), KRAS (47), ARID1A (20), PIK3CA (20) and EGFR (18). There were 38 tier I and 512 tier II variants, with two genes having only a tier I variant, seven genes having both a tier I and tier II variant, and 79 genes having at least one tier II variant. Overall, the ActionSeq™ assay detected clinically significant variants in 42% of the genes included in the panel (88/212), 68% of which (60/88) were detected in more than one tumor type. CONCLUSIONS: This study demonstrates that of the genes with documented involvement in cancer, only a limited number are currently clinically significant from a therapeutic, diagnostic and/or prognostic perspective.

Urolithiasis Is a Risk Factor for Uroseptic Shock and Acute Kidney Injury in Patients With Urinary Tract Infection.

Urinary tract infection (UTI) is a common complication in patients with urolithiasis. This study aimed to compare clinical manifestations and treatment outcomes among UTI patients with or without urolithiasis. It also focused on identifying relationships among urolithiasis, uroseptic shock, and acute kidney injury (AKI). This retrospective study enrolled hospitalized UTI patients who underwent imaging in an acute care setting from January 2006 to March 2015. Of 662 participants enrolled, 113 (17.1%) had urolithiasis, 107 (16.2%) developed uroseptic shock, and 184 (27.8%) developed AKI. A multivariate logistic regression analysis showed that in UTI patients, urolithiasis is associated with an increased risk of uroseptic shock (OR 1.80, 95% CI: 1.08-3.02, P = 0.025), AKI (OR 1.95, 95% CI: 1.22-3.12, P = 0.005), and bacteremia (OR 1.68, 95% CI: 1.08-2.64, P = 0.022). Urolithiasis is common in UTI patients and is associated with an increased risk of uroseptic shock and AKI.

Genotype-Phenotype Correlation in NF1: Evidence for a More Severe Phenotype Associated with Missense Mutations Affecting NF1 Codons 844-848.

Neurofibromatosis type 1 (NF1), a common genetic disorder with a birth incidence of 1:2,000-3,000, is characterized by a highly variable clinical presentation. To date, only two clinically relevant intragenic genotype-phenotype correlations have been reported for NF1 missense mutations affecting p.Arg1809 and a single amino acid deletion p.Met922del. Both variants predispose to a distinct mild NF1 phenotype with neither externally visible cutaneous/plexiform neurofibromas nor other tumors. Here, we report 162 individuals (129 unrelated probands and 33 affected relatives) heterozygous for a constitutional missense mutation affecting one of five neighboring NF1 codons-Leu844, Cys845, Ala846, Leu847, and Gly848-located in the cysteine-serine-rich domain (CSRD). Collectively, these recurrent missense mutations affect ∼0.8% of unrelated NF1 mutation-positive probands in the University of Alabama at Birmingham (UAB) cohort. Major superficial plexiform neurofibromas and symptomatic spinal neurofibromas were more prevalent in these individuals compared with classic NF1-affected cohorts (both p < 0.0001). Nearly half of the individuals had symptomatic or asymptomatic optic pathway gliomas and/or skeletal abnormalities. Additionally, variants in this region seem to confer a high predisposition to develop malignancies compared with the general NF1-affected population (p = 0.0061). Our results demonstrate that these NF1 missense mutations, although located outside the GAP-related domain, may be an important risk factor for a severe presentation. A genotype-phenotype correlation at the NF1 region 844-848 exists and will be valuable in the management and genetic counseling of a significant number of individuals.

Alternative outcomes of pathogenic complex somatic structural variations in the genomes of NF1 and NF2 patients.

Multiplex ligation-dependent probe amplification (MLPA) has been widely used to identify copy-number variations (CNVs), but MLPA's sensitivity and specificity in mosaic CNV detection are largely unknown. Here, we present two mosaic deletions identified by MLPA as NF1 deletion of exons 17-21 and NF2 deletion of exons 9-10. Through cDNA analysis, genomic breakpoint-spanning PCR and Sanger sequencing, we found however both NF1 and NF2 deletions are each composed of two consecutive deletions, which cannot be differentiated by MLPA. Importantly, these consecutive deletions are most likely originating from a single genomic rearrangement and have been preserved independently in different populations of cells.

Risk Factors for Development of Septic Shock in Patients with Urinary Tract Infection.

INTRODUCTION: Severe sepsis and septic shock are associated with substantial mortality. However, few studies have assessed the risk of septic shock among patients who suffered from urinary tract infection (UTI). MATERIALS AND METHODS: This retrospective study recruited UTI cases from an acute care hospital between January 2006 and October 2012 with prospective data collection. RESULTS: Of the 710 participants admitted for UTI, 80 patients (11.3%) had septic shock. The rate of bacteremia is 27.9%; acute kidney injury is 12.7%, and the mortality rate is 0.28%. Multivariable logistic regression analyses indicated that coronary artery disease (CAD) (OR: 2.521, 95% CI: 1.129-5.628, P = 0.024), congestive heart failure (CHF) (OR: 4.638, 95% CI: 1.908-11.273, P = 0.001), and acute kidney injury (AKI) (OR: 2.992, 95% CI: 1.610-5.561, P = 0.001) were independently associated with septic shock in patients admitted with UTI. In addition, congestive heart failure (female, OR: 4.076, 95% CI: 1.355-12.262, P = 0.012; male, OR: 5.676, 95% CI: 1.103-29.220, P = 0.038, resp.) and AKI (female, OR: 2.995, 95% CI: 1.355-6.621, P = 0.007; male, OR: 3.359, 95% CI: 1.158-9.747, P = 0.026, resp.) were significantly associated with risk of septic shock in both gender groups. CONCLUSION: This study showed that patients with a medical history of CAD or CHF have a higher risk of shock when admitted for UTI treatment. AKI, a complication of UTI, was also associated with septic shock. Therefore, prompt and aggressive management is recommended for those with higher risks to prevent subsequent treatment failure in UTI patients.

Risk Factors for Development of Acute Kidney Injury in Patients with Urinary Tract Infection.

Acute kidney injury (AKI) is associated with high morbidity and mortality. Urinary tract infection (UTI) may be associated with sepsis or septic shock, and cause sudden deterioration of renal function. This study investigated the clinical characteristics and change of renal function to identify the risk factors for development of AKI in UTI patients. This retrospective study was conducted in a tertiary referral center. From January 2006 to January 2013, a total of 790 UTI patients necessitating hospital admission were included for final analysis. Their demographic and clinical characteristics and comorbidities were collected and compared. Multivariate logistic regression analysis was performed to evaluate the risk factors for AKI in UTI patients. There were 97 (12.3%) patients developing AKI during hospitalization. Multivariate logistic regression analysis showed that patients with older age (OR 1.02, 95% CI 1.00-1.04, P = 0.04), diabetes mellitus (DM) (OR 2.23, 95% CI 1.35-3.68, P = 0002), upper UTI (OR 2.63, 95% CI 1.53-4.56, P = 0001), afebrile during hospitalization (OR 1.71, 95% CI 1.04-2.83, P = 0036) and lower baseline eGFR [baseline eGFR 45-59 mL/min/1.73 m2 (OR 2.12, 95% CI 1.12-4.04, P = 0.022), baseline eGFR 30-44 mL/min/1.73 m2 (OR 4.44, 95% CI 2.30-8.60 P < 0.001) baseline eGFR < 30 mL/min/1.73 m2 (OR 4.72, 95% CI 2.13-10.45, P <0.001), respectively] were associated with increased risk for development of AKI. were associated with increased risk for development of AKI. Physicians should pay attention to UTI patients at risk of AKI (advancing age, DM, upper UTI, afebrile, and impaired baseline renal function).

Decoding NF1 Intragenic Copy-Number Variations.

Genomic rearrangements can cause both Mendelian and complex disorders. Currently, several major mechanisms causing genomic rearrangements, such as non-allelic homologous recombination (NAHR), non-homologous end joining (NHEJ), fork stalling and template switching (FoSTeS), and microhomology-mediated break-induced replication (MMBIR), have been proposed. However, to what extent these mechanisms contribute to gene-specific pathogenic copy-number variations (CNVs) remains understudied. Furthermore, few studies have resolved these pathogenic alterations at the nucleotide-level. Accordingly, our aim was to explore which mechanisms contribute to a large, unique set of locus-specific non-recurrent genomic rearrangements causing the genetic neurocutaneous disorder neurofibromatosis type 1 (NF1). Through breakpoint-spanning PCR as well as array comparative genomic hybridization, we have identified the breakpoints in 85 unrelated individuals carrying an NF1 intragenic CNV. Furthermore, we characterized the likely rearrangement mechanisms of these 85 CNVs, along with those of two additional previously published NF1 intragenic CNVs. Unlike the most typical recurrent rearrangements mediated by flanking low-copy repeats (LCRs), NF1 intragenic rearrangements vary in size, location, and rearrangement mechanisms. We propose the DNA-replication-based mechanisms comprising both FoSTeS and/or MMBIR and serial replication stalling to be the predominant mechanisms leading to NF1 intragenic CNVs. In addition to the loop within a 197-bp palindrome located in intron 40, four Alu elements located in introns 1, 2, 3, and 50 were also identified as intragenic-rearrangement hotspots within NF1.

Palindrome-mediated and replication-dependent pathogenic structural rearrangements within the NF1 gene.

Palindromic sequences can form hairpin structures or cruciform extrusions, which render them susceptible to genomic rearrangements. A 197-bp long palindromic AT-rich repeat (PATRR17) is located within intron 40 of the neurofibromatosis type 1 (NF1) gene (17q11.2). Through comprehensive NF1 analysis, we identified six unrelated patients with a rearrangement involving intron 40 (five deletions and one reciprocal translocation t(14;17)(q32;q11.2)). We hypothesized that PATRR17 may be involved in these rearrangements thereby causing NF1. Breakpoint cloning revealed that PATRR17 was indeed involved in all of the rearrangements. As microhomology was present at all breakpoint junctions of the deletions identified, and PATRR17 partner breakpoints were located within 7.1 kb upstream of PATRR17, fork stalling and template switching/microhomology-mediated break-induced replication was the most likely rearrangement mechanism. For the reciprocal translocation case, a 51 bp insertion at the translocation breakpoints mapped to a short sequence within PATRR17, proximal to the breakpoint, suggesting a multiple stalling and rereplication process, in contrast to previous studies indicating a purely replication-independent mechanism for PATRR-mediated translocations. In conclusion, we show evidence that PATRR17 is a hotspot for pathogenic intragenic deletions within the NF1 gene and suggest a novel replication-dependent mechanism for PATRR-mediated translocation.