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. 2013 Nov 1;22(21):4339-48.
doi: 10.1093/hmg/ddt283. Epub 2013 Jun 16.

MCTP2 is a dosage-sensitive gene required for cardiac outflow tract development

Seema R Lalani  1 , Stephanie M Ware, Xueqing Wang, Gladys Zapata, Qi Tian, Luis M Franco, Zhengxin Jiang, Kristine Bucasas, Daryl A Scott, Philippe M Campeau, Neil Hanchard, Luis Umaña, Ashley Cast, Ankita Patel, Sau W Cheung, Kim L McBride, Molly Bray, A Craig Chinault, Barbara A Boggs, Miao Huang, Mariah R Baker, Susan Hamilton, Jeff Towbin, John L Jefferies, Susan D Fernbach, Lorraine Potocki, John W Belmont
Affiliations

MCTP2 is a dosage-sensitive gene required for cardiac outflow tract development

Seema R Lalani et al. Hum Mol Genet. .

Abstract

Coarctation of the aorta (CoA) and hypoplastic left heart syndrome (HLHS) have been reported in rare individuals with large terminal deletions of chromosome 15q26. However, no single gene important for left ventricular outflow tract (LVOT) development has been identified in this region. Using array-comparative genomic hybridization, we identified two half-siblings with CoA with a 2.2 Mb deletion on 15q26.2, inherited from their mother, who was mosaic for this deletion. This interval contains an evolutionary conserved, protein-coding gene, MCTP2 (multiple C2-domains with two transmembrane regions 2). Using gene-specific array screening in 146 individuals with non-syndromic LVOT obstructive defects, another individual with HLHS and CoA was found to have a de novo 41 kb intragenic duplication within MCTP2, predicted to result in premature truncation, p.F697X. Alteration of Mctp2 gene expression in Xenopus laevis embryos by morpholino knockdown and mRNA overexpression resulted in the failure of proper OT development, confirming the functional importance of this dosage-sensitive gene for cardiogenesis. Our results identify MCTP2 as a novel genetic cause of CoA and related cardiac malformations.

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Figures

Figure 1.
Figure 1.
Submicroscopic deletion of MCTP2 within 15q26.2 in two half-siblings with CoA. (A  and B) Subject 1 and his half-sister, subject 2 (C and D) with pertinent features of CoA, microbrachycephaly, malar hypoplasia, thin and down-turned upper lip, posteriorly rotated ears and short stature. (E) SNP genotyping using Illumina HumanHap300 BeadChip array showing a 2.2 Mb deletion involving a single gene, MCTP2 on 15q26.2 in subject 1. (F) Genomic loss of copy-number detected by RP11-4F5 in subject 1 showing loss of hybridization signal (red) on one copy of chromosome 15 (shown by arrow). (G) These results were also confirmed in his half-sister. Delineation of mosaicism for this deletion in their mother with 26% of her cells showing normal hybridization signals (H) and 74% of the cells showing a single hybridization signal for RP11-4F5 (I).
Figure 2.
Figure 2.
MCTP2-specific targeted array-CGH analysis in an individual with HLHS with CoA and breakpoint mapping. (A) A 41 kb de novo intragenic duplication of MCTP2 was identified in patient 3 on the targeted array-CGH and was confirmed by interphase fluorescence in situ hybridization (FISH) analysis by fosmid (B, highlighted by the arrow). (C) A junction-specific PCR amplification product was amplified from DNA from patient 3 ‘P’, but not in a control individual ‘C’, his father ‘F’ or his mother ‘M’. (D) DNA sequence from the junction fragment indicating that the breakpoints of the 41 kb duplication lie within intron 17 and intron 10 of the MCTP2 gene. (E and F) A schematic representation of the resulting mRNA product with the duplicated fragment is show in red. If correctly spliced, this mRNA is predicted to encode a truncated protein, p.F697X.
Figure 3.
Figure 3.
Mctp2 morphant analyses. (A) Embryos were scored for viability at the indicated stages. Sub-effective doses of morpholinos had no effect on phenotype or viability when compared with controls when injected individually [Ctrl, MO1 1:50 (0.54 ng), MO2 1:50 (0.54 ng)]. Morphants injected with both morpholinos at sub-effective doses showed significant decreases in viability. The number of embryos analyzed in each group is represented above each group of bars. (B–D) Control and Mctp2 morphant embryo morphology at stage 44/45. Qualitatively similar phenotypic results were achieved using both morpholinos and (C) and (D) are representative images using MO1. (C) Mctp2 morphants have grossly normal external morphology with the exception of moderate to severe edema in a subset of embryos (arrows). (D) High power view of embryo with severe edema showing accumulation of fluid (arrowhead) in the cardiac region (arrow at heart).
Figure 4.
Figure 4.
Altering Mctp2 expression levels causes defects in cardiac development. (A) Comparison of rate of cardiac defects in morphants injected at sub-effective morpholino concentrations singly or in combination. (B and C) Whole-mount in situ hybridization with cardiac troponin T illustrates abnormal cardiac morphology in MO1/MO2 morphant (C) when compared with control (B). Representative histological analyses of control (D and F) and Mctp2 MO1 (E and G) embryos at stage 44/45 demonstrate abnormalities of the OT and endocardial cushion formation. (D and E) Posterior/proximal OT at the level of connection with the ventricle. (F and G) High powered views of the developing OT in (D) and (E) are shown. In control embryos, the endocardial cushion of the developing spiral valve fills the OT throughout its proximal to distal length. In contrast, sections of Mctp2 morphant embryos showed no evidence of endocardial cushion formation at any level of the developing OT, confirming the critical functional role of Mctp2 in cardiac OT development. (H) Percent of embryos with abnormal OT development. Transverse sections were analyzed for Mctp2 MO1, Mctp2 MO2 and Mctp2 mRNA-injected embryos. The number of embryos analyzed for each injection group is indicated above the bar. a, atria; cj, cardiac jelly; e, endocardial cells; ec, endocardial cushion; oft, outflow tract; V, ventricle.
Figure 5.
Figure 5.
Ca2+ titration of the terbium fluorescence showing altered Ca2+-binding affinity of the C2A domain mutants. Terbium fluorescence was used to monitor the changes in relative affinity of Ca2+ for the C2A domain. Terbium(III) (1 mm) was incubated with MCTP2-C2A (20 µm) and titrated with Ca2+ (5 nm–50 mm). The terbium(III) signal (excitation 285 nm; emission 545 nm) decreased with increasing Ca2+ concentrations and the data are plotted as the percent decrease from 0 Ca2+. The IC50 s for the wild-type C2A are 524 ± 2 nm and 28 ± 1 µm; for G203D are 61 ± 1 nm (P < 0.0001 compared with wild-type site 1) and 99 ± 1 µm (P < 0.0001 compared with wild-type site 2) and for Y235C are 05 ± 1 nm (P < 0.0001 compared with wild-type site 1) and 35 ± 1 µm (P = 0.0173 compared with wild-type site 2). Data shown are means ± SEM (n = 3–6).
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References

    1. Hoffman J.I., Kaplan S. The incidence of congenital heart disease. J. Am. Coll. Cardiol. 2002;39:1890–1900. - PubMed
    1. Boughman J.A., Berg K.A., Astemborski J.A., Clark E.B., McCarter R.J., Rubin J.D., Ferencz C. Familial risks of congenital heart defect assessed in a population-based epidemiologic study. Am. J. Med. Genet. A. 1987;26:839–849. - PubMed
    1. Ferencz C., Boughman J.A. Congenital heart disease in adolescents and adults. teratology, genetics, and recurrence risks. Cardiol. Clin. 1993;11:557–567. - PubMed
    1. Gerboni S., Sabatino G., Mingarelli R., Dallapiccola B. Coarctation of the aorta, interrupted aortic arch, and hypoplastic left heart syndrome in three generations. J. Med. Genet. 1993;30:328–329. - PMC - PubMed
    1. Wessels M.W., Berger R.M., Frohn-Mulder I.M., Roos-Hesselink J.W., Hoogeboom J.J., Mancini G.S., Bartelings M.M., Krijger R., Wladimiroff J.W., Niermeijer M.F., et al. Autosomal dominant inheritance of left ventricular outflow tract obstruction. Am. J. Med. Genet. A. 2005;134:171–179. - PubMed

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