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. 2024 Dec 18;16(1):146.
doi: 10.1186/s13073-024-01411-7.

Structural variant allelic heterogeneity in MECP2 duplication syndrome provides insight into clinical severity and variability of disease expression

Davut Pehlivan #  1   2   3   4 , Jesse D Bengtsson #  5 , Sameer S Bajikar #  6   7 , Christopher M Grochowski  6   8 , Ming Yin Lun  5 , Mira Gandhi  5 , Angad Jolly  6 , Alexander J Trostle  9   7 , Holly K Harris  9   10 , Bernhard Suter  11   9 , Sukru Aras  7 , Melissa B Ramocki  11   12 , Haowei Du  6 , Michele G Mehaffey  5 , KyungHee Park  5 , Ellen Wilkey  5 , Cemal Karakas  13 , Jesper J Eisfeldt  14 , Maria Pettersson  14 , Lynn Liu  15 , Marwan S Shinawi  16 , Virginia E Kimonis  17 , Wojciech Wiszniewski  18 , Kyle Mckenzie  19 , Timo Roser  20 , Angela M Vianna-Morgante  21 , Alberto S Cornier  22 , Ahmed Abdelmoity  23 , James P Hwang  8 , Shalini N Jhangiani  8 , Donna M Muzny  8 , Tadahiro Mitani  24 , Kazuhiro Muramatsu  24 , Shin Nabatame  25 , Daniel G Glaze  11   9 , Jawid M Fatih  6 , Richard A Gibbs  6   8 , Zhandong Liu  9   7 , Anna Lindstrand  14 , Fritz J Sedlazeck  8 , James R Lupski  6   9   8   26 , Huda Y Zoghbi  27   28   29   30   31 , Claudia M B Carvalho  32   33
Affiliations

Structural variant allelic heterogeneity in MECP2 duplication syndrome provides insight into clinical severity and variability of disease expression

Davut Pehlivan et al. Genome Med. .

Abstract

Background: MECP2 Duplication Syndrome, also known as X-linked intellectual developmental disorder Lubs type (MRXSL; MIM: 300260), is a neurodevelopmental disorder caused by copy number gains spanning MECP2. Despite varying genomic rearrangement structures, including duplications and triplications, and a wide range of duplication sizes, no clear correlation exists between DNA rearrangement and clinical features. We had previously demonstrated that up to 38% of MRXSL families are characterized by complex genomic rearrangements (CGRs) of intermediate complexity (2 ≤ copy number variant breakpoints < 5), yet the impact of these genomic structures on regulation of gene expression and phenotypic manifestations have not been investigated.

Methods: To study the role of the genomic rearrangement structures on an individual's clinical phenotypic variability, we employed a comprehensive genomics, transcriptomics, and deep phenotyping analysis approach on 137 individuals affected by MRXSL. Genomic structural information was correlated with transcriptomic and quantitative phenotypic analysis using Human Phenotype Ontology (HPO) semantic similarity scores.

Results: Duplication sizes in the cohort ranging from 64.6 kb to 16.5 Mb were classified into four categories comprising of tandem duplications (48%), terminal duplications (22%), inverted triplications (20%), and other CGRs (10%). Most of the terminal duplication structures consist of translocations (65%) followed by recombinant chromosomes (23%). Notably, 65% of de novo events occurred in the Terminal duplication group in contrast with 17% observed in Tandem duplications. RNA-seq data from lymphoblastoid cell lines indicated that the MECP2 transcript quantity in MECP2 triplications is statistically different from all duplications, but not between other classes of genomic structures. We also observed a significant (p < 0.05) correlation (Pearson R = 0.6, Spearman p = 0.63) between the log-transformed MECP2 RNA levels and MECP2 protein levels, demonstrating that genomic aberrations spanning MECP2 lead to altered MECP2 RNA and MECP2 protein levels. Genotype-phenotype analyses indicated a gradual worsening of phenotypic features, including overall survival, developmental levels, microcephaly, epilepsy, and genitourinary/eye abnormalities in the following order: Tandem duplications, Other complex duplications, Terminal duplications/Translocations, and Triplications encompassing MECP2.

Conclusion: In aggregate, this combined analysis uncovers an interplay between MECP2 dosage, genomic rearrangement structure and phenotypic traits. Whereas the level of MECP2 is a key determinant of the phenotype, the DNA rearrangement structure can contribute to clinical severity and disease expression variability. Employing this type of analytical approach will advance our understanding of the impact of genomic rearrangements on genomic disorders and may help guide more targeted therapeutic approaches.

Keywords: MECP2 duplication syndrome; Clinical severity; MRXSL; Survival; Tandem duplication; Terminal duplication.

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Conflict of interest statement

Declarations. Ethics approval and consent to participate: This study is approved by Baylor College of Medicine (BCM) and Pacific Northwest Research Institute (PNRI). For patients who were evaluated at TCH-BBC Rett Center, clinical information was obtained by retrospective chart review using H-46044 protocol approved by Baylor College of Medicine’s (BCM) Institutional Review Board (IRB). We used BCM IRB approved H-32407 and H-47281/PNRI WIRB #20202158 protocols to clinically examine patients on a research basis. For genomic studies, participants were consented according to the IRB at BCM approved protocols: H-29697, H-20268, H-18122, and H-26667 or H-47281/Pacific Northwest Research Institute WIRB #20202158. The research was conducted in compliance with the principles of Helsinki Declaration. Consent for publication: All subjects gave consent for participation into our study and publication of genomic and clinical information. Competing interests: BCM and Miraca Holdings have formed a joint venture with shared ownership and governance of BG, which performs clinical microarray analysis (CMA), clinical ES (cES), and clinical biochemical studies. J.R.L. serves on the Scientific Advisory Board of the BG. J.R.L. has stock ownership in 23andMe, is a paid consultant for Genomics International, and is a coinventor on multiple United States and European patents related to molecular diagnostics for inherited neuropathies, eye diseases, genomic disorders and bacterial genomic fingerprinting. D.P. and M.B.R. provide consulting service for Ionis Pharmaceuticals. The remaining authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
Distribution of genomic structures observed in the MRXSL cohort. a Pie chart distribution of the genomic structure of 118 unrelated individuals carrying MECP2 duplication. b Violin plot representing the size distribution of the CNV encompassing MECP2 in each genomic subgroup. Length in bp, log10 scale. c Pie chart distribution of the genomic structure of 23 de novo MECP2 duplication events. d Violin plot representing the size distribution of inherited and de novo structural variants in MRXSL. CGR: Complex Genomic Rearrangement, DUP: Duplication, INV: Inverted, NML: Normal, rec: Recombinant, SV: Structural Variant, TRP: Triplication
Fig. 2
Fig. 2
Developmental Comparisons of Different MRXSL structural variants. Highest achieved gross motor skills (a), gross motor DQ (b), highest achieved fine motor skills (c), fine motor DQ (d). Overall, the developmental delay severity worsens in the following order: Tandem duplication < Other complex duplication < Terminal duplication < Translocation < Triplication. Medians are provided in gray boxes/black lines, means are given in blue lines. MRXSL: MECP2 Duplication Syndrome, DQ: Developmental Quotient
Fig. 3
Fig. 3
Survival curve analysis for different genomic subgroups. Cox regression for survival analysis indicate that SVs are good predictor of survival probability: Tandem (5/59) > Other complex (3/41) > Translocations (4/16) > Triplications (4/5) (Additional file: Table S1). Overall p-value < 0.0001. SV: Structural Variant
Fig. 4
Fig. 4
Transcriptomic heterogeneity amongst MRXSL patient-derived lymphoblastoid cell lines. RNA from lymphoblasts from 64 MRXSL individuals from the clinical cohort was collected and processed for RNA-sequencing transcriptomic analyses. Patient lines were collected as triplicate RNA preparations. a Normalized MECP2 expression from RNA-sequencing data. b Normalized IRAK1 expression from RNA-sequencing data. c Gene expression along Xq28. Expressed genes are ordered from more centromeric (left) to telomeric (right) within the maximal genomic region spanning the cohort of samples collected. Samples are scaled by column, and rows are clustered using hierarchical clustering using Euclidean distance. d Principal component representation of global gene expression changes between MRXSL and unaffected control lines
Fig. 5
Fig. 5
Correlation between MECP2 RNA and MECP2 protein measurements in patient LCLs. a MECP2 protein measurements were made using capillary electrophoresis from matched lysates as collected for RNA-sequencing. MECP2 signal intensity was measured using a Jess Western instrument and normalized to total protein. b Correlation between MECP2 RNA and MECP2 protein; Pearson and Spearman correlation coefficients are displayed in upper left
Fig. 6
Fig. 6
Phenotypic analysis of five different structural variant groups of MRXSL and quantitative similarity analysis of MRXSL features with known OMIM genes on the p and q terminals of X-chromosome. a Prevalence of certain features ranged from 0 (light yellow) to 1.0 (black). Individuals carrying Tandem Duplication and Other complex duplication, Terminal duplication and Translocation have similar patterns. Individuals with MECP2 triplication pattern is different than the other four groups. The scale is provided on the right. DUP: Duplication, TRP: Triplication. b The heatmap analysis showed that the highest overlap is among the different structural variants of MRXSL, followed by MECP2, FMR1, SLC6A8, NAA10 and FLNA. Of note, six individuals were excluded from HPO analysis since there were either too little or too many HPO terms
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