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Dubowitz syndrome is a complex comprised of multiple, genetically distinct and phenotypically overlapping disorders.

Stewart DR, Pemov A, Johnston JJ, Sapp JC, Yeager M, He J, Boland JF, Burdett L, Brown C, Gatti RA, Alter BP, Biesecker LG, Savage SA - PLoS ONE (2014)

Bottom Line: Western blotting showed an absence of a ligase IV band in both siblings.In the third patient, array SNP genotyping revealed a de novo ∼ 3.89 Mb interstitial deletion at chromosome 17q24.2 (chr 17:62,068,463-65,963,102, hg18), which spanned the known Carney complex gene PRKAR1A.Our work suggests that, in addition to dyskeratosis congenita, LIG4 and 17q24.2 syndromes also feature shortened telomeres; to confirm this, telomere length testing should be considered in both disorders.

View Article: PubMed Central - PubMed

Affiliation: Clinical Genetics Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, NIH, Rockville, Maryland, United States of America.

ABSTRACT
Dubowitz syndrome is a rare disorder characterized by multiple congenital anomalies, cognitive delay, growth failure, an immune defect, and an increased risk of blood dyscrasia and malignancy. There is considerable phenotypic variability, suggesting genetic heterogeneity. We clinically characterized and performed exome sequencing and high-density array SNP genotyping on three individuals with Dubowitz syndrome, including a pair of previously-described siblings (Patients 1 and 2, brother and sister) and an unpublished patient (Patient 3). Given the siblings' history of bone marrow abnormalities, we also evaluated telomere length and performed radiosensitivity assays. In the siblings, exome sequencing identified compound heterozygosity for a known rare nonsense substitution in the nuclear ligase gene LIG4 (rs104894419, NM_002312.3:c.2440C>T) that predicts p.Arg814X (MAF:0.0002) and an NM_002312.3:c.613delT variant that predicts a p.Ser205Leufs*29 frameshift. The frameshift mutation has not been reported in 1000 Genomes, ESP, or ClinSeq. These LIG4 mutations were previously reported in the sibling sister; her brother had not been previously tested. Western blotting showed an absence of a ligase IV band in both siblings. In the third patient, array SNP genotyping revealed a de novo ∼ 3.89 Mb interstitial deletion at chromosome 17q24.2 (chr 17:62,068,463-65,963,102, hg18), which spanned the known Carney complex gene PRKAR1A. In all three patients, a median lymphocyte telomere length of ≤ 1st centile was observed and radiosensitivity assays showed increased sensitivity to ionizing radiation. Our work suggests that, in addition to dyskeratosis congenita, LIG4 and 17q24.2 syndromes also feature shortened telomeres; to confirm this, telomere length testing should be considered in both disorders. Taken together, our work and other reports on Dubowitz syndrome, as currently recognized, suggest that it is not a unitary entity but instead a collection of phenotypically similar disorders. As a clinical entity, Dubowitz syndrome will need continual re-evaluation and re-definition as its constituent phenotypes are determined.

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Neutral comet assay compatible with a DNA double-strand break repair disorder.(a). Tail moment of WT, A-T and patient cells at 0, 30 minutes and 5 hours post-irradiation. All cells show damage at 30 minutes post-irradiation. WT returns to near baseline levels after 5 hours, while A-T and patient cells show long comet tails after 5 hours, indicating significant levels of unrepaired DNA. (b). Ratio of unrepaired DNA at 5/0 hours. A-T as well as Patient 2 and 3 cells show statistically significantly lower levels of repair at 5 hours post-irradiation compared to wild type cells (* p<0.05). Experiments were performed in triplicate with the average of three experiments shown. WT  =  wild type; A-T  =  ataxia-telangiectasia control.
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pone-0098686-g009: Neutral comet assay compatible with a DNA double-strand break repair disorder.(a). Tail moment of WT, A-T and patient cells at 0, 30 minutes and 5 hours post-irradiation. All cells show damage at 30 minutes post-irradiation. WT returns to near baseline levels after 5 hours, while A-T and patient cells show long comet tails after 5 hours, indicating significant levels of unrepaired DNA. (b). Ratio of unrepaired DNA at 5/0 hours. A-T as well as Patient 2 and 3 cells show statistically significantly lower levels of repair at 5 hours post-irradiation compared to wild type cells (* p<0.05). Experiments were performed in triplicate with the average of three experiments shown. WT  =  wild type; A-T  =  ataxia-telangiectasia control.

Mentions: The neutral comet assay measures the level of unrepaired DNA in a cell [14]–[16]. Recent studies have shown a correlation between double-strand DNA post-irradiation damage, as measured by NCA, and the colony survival assay [14]. Longer, undamaged DNA fragments will remain predominantly in the “head” of the comet, while smaller, heavily damaged DNA fragments will migrate more quickly and form the tail of the comet. The average tail moment at 5 hours post-irradiation is divided by the average tail moment of non-irradiated cells to determine the percent DNA repair. Wild type cells with normal DNA double-strand-break repair abilities will re-ligate most breaks within 5 hours while cells with deficient repair mechanisms will have a much lower percent DNA repair. The results from both patients 2 and 3 show large amounts of unrepaired DNA at 5 hours post-irradiation (Figure 9). Their percent repair is significantly less than wild type cells and is similar to the repair efficiency of ATM-deficient cells. These results, along with the concordant, abnormal CSA scores in both patients are compatible with a DNA double-strand break repair disorder.


Dubowitz syndrome is a complex comprised of multiple, genetically distinct and phenotypically overlapping disorders.

Stewart DR, Pemov A, Johnston JJ, Sapp JC, Yeager M, He J, Boland JF, Burdett L, Brown C, Gatti RA, Alter BP, Biesecker LG, Savage SA - PLoS ONE (2014)

Neutral comet assay compatible with a DNA double-strand break repair disorder.(a). Tail moment of WT, A-T and patient cells at 0, 30 minutes and 5 hours post-irradiation. All cells show damage at 30 minutes post-irradiation. WT returns to near baseline levels after 5 hours, while A-T and patient cells show long comet tails after 5 hours, indicating significant levels of unrepaired DNA. (b). Ratio of unrepaired DNA at 5/0 hours. A-T as well as Patient 2 and 3 cells show statistically significantly lower levels of repair at 5 hours post-irradiation compared to wild type cells (* p<0.05). Experiments were performed in triplicate with the average of three experiments shown. WT  =  wild type; A-T  =  ataxia-telangiectasia control.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4043752&req=5

pone-0098686-g009: Neutral comet assay compatible with a DNA double-strand break repair disorder.(a). Tail moment of WT, A-T and patient cells at 0, 30 minutes and 5 hours post-irradiation. All cells show damage at 30 minutes post-irradiation. WT returns to near baseline levels after 5 hours, while A-T and patient cells show long comet tails after 5 hours, indicating significant levels of unrepaired DNA. (b). Ratio of unrepaired DNA at 5/0 hours. A-T as well as Patient 2 and 3 cells show statistically significantly lower levels of repair at 5 hours post-irradiation compared to wild type cells (* p<0.05). Experiments were performed in triplicate with the average of three experiments shown. WT  =  wild type; A-T  =  ataxia-telangiectasia control.
Mentions: The neutral comet assay measures the level of unrepaired DNA in a cell [14]–[16]. Recent studies have shown a correlation between double-strand DNA post-irradiation damage, as measured by NCA, and the colony survival assay [14]. Longer, undamaged DNA fragments will remain predominantly in the “head” of the comet, while smaller, heavily damaged DNA fragments will migrate more quickly and form the tail of the comet. The average tail moment at 5 hours post-irradiation is divided by the average tail moment of non-irradiated cells to determine the percent DNA repair. Wild type cells with normal DNA double-strand-break repair abilities will re-ligate most breaks within 5 hours while cells with deficient repair mechanisms will have a much lower percent DNA repair. The results from both patients 2 and 3 show large amounts of unrepaired DNA at 5 hours post-irradiation (Figure 9). Their percent repair is significantly less than wild type cells and is similar to the repair efficiency of ATM-deficient cells. These results, along with the concordant, abnormal CSA scores in both patients are compatible with a DNA double-strand break repair disorder.

Bottom Line: Western blotting showed an absence of a ligase IV band in both siblings.In the third patient, array SNP genotyping revealed a de novo ∼ 3.89 Mb interstitial deletion at chromosome 17q24.2 (chr 17:62,068,463-65,963,102, hg18), which spanned the known Carney complex gene PRKAR1A.Our work suggests that, in addition to dyskeratosis congenita, LIG4 and 17q24.2 syndromes also feature shortened telomeres; to confirm this, telomere length testing should be considered in both disorders.

View Article: PubMed Central - PubMed

Affiliation: Clinical Genetics Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, NIH, Rockville, Maryland, United States of America.

ABSTRACT
Dubowitz syndrome is a rare disorder characterized by multiple congenital anomalies, cognitive delay, growth failure, an immune defect, and an increased risk of blood dyscrasia and malignancy. There is considerable phenotypic variability, suggesting genetic heterogeneity. We clinically characterized and performed exome sequencing and high-density array SNP genotyping on three individuals with Dubowitz syndrome, including a pair of previously-described siblings (Patients 1 and 2, brother and sister) and an unpublished patient (Patient 3). Given the siblings' history of bone marrow abnormalities, we also evaluated telomere length and performed radiosensitivity assays. In the siblings, exome sequencing identified compound heterozygosity for a known rare nonsense substitution in the nuclear ligase gene LIG4 (rs104894419, NM_002312.3:c.2440C>T) that predicts p.Arg814X (MAF:0.0002) and an NM_002312.3:c.613delT variant that predicts a p.Ser205Leufs*29 frameshift. The frameshift mutation has not been reported in 1000 Genomes, ESP, or ClinSeq. These LIG4 mutations were previously reported in the sibling sister; her brother had not been previously tested. Western blotting showed an absence of a ligase IV band in both siblings. In the third patient, array SNP genotyping revealed a de novo ∼ 3.89 Mb interstitial deletion at chromosome 17q24.2 (chr 17:62,068,463-65,963,102, hg18), which spanned the known Carney complex gene PRKAR1A. In all three patients, a median lymphocyte telomere length of ≤ 1st centile was observed and radiosensitivity assays showed increased sensitivity to ionizing radiation. Our work suggests that, in addition to dyskeratosis congenita, LIG4 and 17q24.2 syndromes also feature shortened telomeres; to confirm this, telomere length testing should be considered in both disorders. Taken together, our work and other reports on Dubowitz syndrome, as currently recognized, suggest that it is not a unitary entity but instead a collection of phenotypically similar disorders. As a clinical entity, Dubowitz syndrome will need continual re-evaluation and re-definition as its constituent phenotypes are determined.

Show MeSH
Related in: MedlinePlus