Intragenic MBD5 familial deletion variant does not negatively impact MBD5 mRNA expression
© Mullegama and Elsea; licensee BioMed Central Ltd. 2014
Received: 18 September 2014
Accepted: 25 October 2014
Published: 19 November 2014
2q23.1 deletion syndrome is characterized by intellectual disability, speech impairment, seizures, disturbed sleep pattern, behavioral problems, and hypotonia. Core features of this syndrome are due to haploinsufficiency of MBD5. Deletions that include coding and noncoding exons show reduced MBD5 mRNA expression. We report a patient with a neurological and behavioral phenotype similar to 2q23.1 deletion syndrome with an inherited intronic deletion in the 5-prime untranslated region of MBD5. Our data show that this patient has normal MBD5 mRNA expression; therefore, this deletion is likely not causative for 2q23.1 deletion syndrome. Overall, it is important to validate intronic deletions for pathogenicity.
Comparison of the phenotype of SMS431 to the prominent features of 2q23.1 deletion syndrome
2q23.1 deletion syndrome
Repetitive behaviors (stereotypies)
Short attention span
Infantile feeding difficulties
Heavy arched eyebrows
Thin upper lip
Widely spaced teeth
The Institutional Review Board Baylor College of Medicine approved this study. Fresh blood was collected from SMS431 (proband), SMS432 (father), SMS433 (mother), SMS361 (2q23.1 deletion patient) , and nine normal controls after informed consent was obtained. Total RNA was isolated according to standard methods (Invitrogen, Carlsbad, CA). RNA was quantified using the NanoDrop ® ND-100 Spectrophotometer and reverse transcribed through qSCRIPT cDNA SuperMix (Quanta Biosciences, Inc., Gaithersburg, MD) according to manufacturer’s instructions. To assess MBD5 mRNA expression, quantitative RT-PCR was performed as previously described ,. Briefly, Taqman minor groove binder probes for MBD5 (OMIM 611472, Hs00289233_m1) and GAPDH (OMIM 138400, Hs9999905_m1) were used. GAPDH was used as the endogenous control. All samples of cDNA were run in triplicate in 10 ul reaction volumes. All samples were run and analyzed according to previously published methods using BioRad CFX Connect™ Real-Time PCR Detection System . Three biological replicates were performed. Results are expressed as fold-change relative to the control sample. Standard error was generated for each sample. A paired t- test was used to determine significance. p <0.01 was considered statistically significant.
A survey of the Database of Genomic Variants (DGV) contains no deletions that are identical to that observed in the individuals in this family. DVG lists 14 small deletions in the 5′-noncoding region of MBD5, with nine deletions confined to intronic sequences, which were reported to be nonpathogenic or of unknown significance. Five of the nine small deletions were found in intron 4.
Overall, it is apparent that the intron 4 deletion does not affect expression of MBD5 and likely does not give rise to the features associated with the MBD5 haploinsufficiency that is observed in 2q23.1 deletion syndrome.
Introns in the noncoding region of a gene can play a major role in the transcriptional regulation of a gene and consequently, gene expression. An intron can enhance gene expression through the presence of transcriptional regulatory elements or through structural modulation and splicing . Copy number variants within splice site sequences at the intron-exon junction cause approximately 10% of disease-causing mutations . There are several cases in the medical literature of pathogenic intronic deletions such as NRXN1 deletions (Autism Spectrum Disorder) , SLC34A3 deletions (hereditary hypophosphatemic rickets with hypercalciuria) ,, PKD1 deletions (Rothmund-Thomson syndrome) , and NASE deletions (5-fluorouracil toxicity) .
Since the intronic deletion identified in SMS431 does not delete the splice junctions between exon 4 and exon 5 or create new splice junctions, proper mRNA splicing likely still occurs, without impacting expression. Further, deletions in 5-prime UTR introns could lead to altered expression if transcription factor binding is affected. Whether there are transcription factors, enhancers, or silencers that bind to that specific intronic region of MBD5 remains unclear.
In summary, SMS431 exhibited developmental delay, motor delay, severe language impairment, sleep disturbances and behavioral problems that mimicked 2q23.1 deletion syndrome (Table 1). However, these neurological and behavioral phenotypes are observed in other neurodevelopmental disorders. The 2q23.1 deletion syndrome phenotype overlaps with other neurodevelopmental disorders, including Angelman syndrome, Smith-Magenis syndrome, Pitt-Hopkins syndrome, and Kleefstra syndrome ,. SMS431 likely has a different neurodevelopmental disorder that overlaps phenotypically with 2q23.1 deletion syndrome. Since no other copy number variants were identified for this patient, additional testing will be required to determine a cause for the phenotype. The possibility exists that a gene that is involved in a pathway common to MBD5 may be affected, including a gene downstream of MBD5 or binding partner of MBD5. Finally, whole exome sequencing (WES) may be the best approach toward diagnosis for this child.
In conclusion, we highlight the importance of validating intronic deletions for pathogenicity so that accurate and informed diagnosis can be provided to the patient.
Written informed consent was obtained from the patient’s parents for publication of this paper. A copy of the written consent is available for review by the Editor-in-Chief of this journal.
We are thankful to the family for their willingness to participate in this study. We thank the Fondation Jerome Lejuene for funding portions of this study. This work was supported in part by resources from the Baylor College of Medicine and the Jan and Dan Duncan Neurological Research Institute.
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