Down syndrome is a condition caused by having an extra copy of one of the 46 chromosomes found inside human cells. Specifically, instead of two copies, people with Down syndrome are born with three copies of chromosome 21. This results in many different effects, including learning and memory problems, heart defects and Alzheimer’s disease. Each of these different effects is caused by having a third copy of one or more of the approximately 230 genes found on chromosome 21. However, it is not known which of these genes cause any of these effects, and how an extra copy of the genes results in such changes.

Now, Lana-Elola et al. have investigated which genes on chromosome 21 cause the heart defects seen in Down syndrome, and how those heart defects come about. This involved engineering a new strain of mouse that has an extra copy of 148 mouse genes that are very similar to 148 genes found on chromosome 21 in humans. Like people with Down syndrome, this mouse strain developed heart defects when it was an embryo.

Using a series of six further mouse strains, Lana-Elola et al. then narrowed down the potential genes that, when in three copies, are needed to cause the heart defects, to a list of just 39 genes. Further experiments then showed that at least two genes within these 39 genes were required in three copies to cause the heart defects.

The next step will be to identify the specific genes that actually cause the heart defects, and then work out how a third copy of these genes causes the developmental problems.

The formation of a functional four-chambered heart is a complex process and perturbations in its development can lead to congenital heart defects (CHD). These affect almost 1% of the population and are a major cause of morbidity and infant mortality (Fahed et al., 2013). The chance of being born with CHD is drastically increased (to ~50%) in DS (Vis et al., 2009) in which a range of heart defects is seen, including ventricular septal defects (VSD) and outflow tract abnormalities such as overriding aorta (OA) and double outlet right ventricle (DORV). Notably, defects that affect the atrioventricular (AV) junction, especially atrio-ventricular septal defects (AVSD) (Freeman et al., 1998) are often seen in DS. AVSD comprise a spectrum of cardiac malformations characterized by a common AV junction, guarded by an essentially common valve, as opposed to separate AV junctions guarded by mitral and tricuspid valves. People diagnosed with AVSD can present with communication between left and right heart chambers (shunting) at the atrial level (ostium primum defect) or at the ventricular level or with shunting at both atrial and ventricular levels (Mahle et al., 2006).

The process of forming the AV junction that divides the embryonic heart into a four-chambered structure involves the growth and fusion of a number of tissues from different precursor populations (Webb et al., 1998). These include the atrial and ventricular septa (mainly of myocardial origin), the mesenchymal cap of the atrial septum, and the superior and inferior endocardial cushions (endocardial origin). In addition, the dorsal mesenchymal protrusion (DMP) first described as the ‘spina vestibuli’ by Wilhelm His the elder in 1880 (His, 1880; Mommersteeg et al., 2006) makes an important contribution to septation of the AV junction (Anderson et al., 2015; Snarr et al., 2007a; Snarr et al., 2007b; Webb et al., 1998). The DMP is a mesenchymal structure at the venous pole of the developing heart derived from the second heart field (SHF) and perturbations of its development result in AVSD (Briggs et al., 2013; Goddeeris et al., 2008; Rana et al., 2014; Tian et al., 2010; Webb et al., 1999; Xie et al., 2012). Abnormalities in the DMP have also been detected in human fetal hearts with trisomy 21, leading to the hypothesis that malformation of this tissue causes the AVSD in DS (Blom et al., 2003).

Hsa21 carries 233 protein-coding genes (genome assembly GRCh38.p5) and it is thought that DS phenotypes result from an increased dosage of one or more of the genes on Hsa21. The search for dosage-sensitive genes that when present in 3 copies cause DS phenotypes has been approached using both human and mouse genetics. In humans, rare cases of partial trisomy 21 have been used to identify critical regions that contain dosage-sensitive genes that when present in 3 copies cause DS phenotypes (Delabar et al., 1993; Korbel et al., 2009; Korenberg et al., 1994; Lyle et al., 2009). Alternatively, mouse strains have been generated to study the pathology and genetics of DS.

Hsa21 shares synteny with a large region on mouse chromosome 16 (Mmu16) and with shorter regions on Mmu10 and Mmu17. The first two DS models generated, Ts65Dn (Davisson et al., 1990) and Ts1Cje (Sago et al., 1998) have duplications of regions of mouse chromosome 16 (Mmu16) that are orthologous to Hsa21, and have been used to identify dosage-sensitive genes contributing to some DS phenotypes (Baek et al., 2009; Chakrabarti et al., 2010; Lana-Elola et al., 2011; Salehi et al., 2006; Sussan et al., 2008). However, both models also have additional aneuploidy (trisomy of 60 genes on Mmu17 in Ts65Dn and monosomy of 7 genes on Mmu12 in Ts1Cje mice), making interpretation of their phenotypes difficult (Duchon et al., 2011). Another DS model, the Tc1 mouse, carries a freely-segregating copy of Hsa21 and shows many DS phenotypes, including CHD (Dunlevy et al., 2010; O'Doherty et al., 2005), although the Hsa21 in this strain is not intact and the mice are mosaic for the human chromosome, again making interpretation difficult (Gribble et al., 2013). Thanks to recent advances in chromosome engineering, a number of mouse strains with duplications of regions of mouse chromosomes orthologous to Hsa21 have been generated, resulting in partial trisomies, including the most complete mouse model for DS to date (Dp(16)1Yey/+;Dp(17)1Yey/+;Dp(10)1Yey/+), which carries a duplication of all Hsa21-orthologous regions on Mmu16, Mmu17 and Mmu10 (Brault et al., 2015; Li et al., 2007; Liu et al., 2013; 2011; Olson et al., 2004; Pereira et al., 2009; Yu et al., 2010). However, a comprehensive mapping panel to finely map dosage-sensitive genes in DS has not been available until now.

To identify dosage-sensitive critical regions and candidate genes, we now report the generation of a fine mapping panel of 7 partial trisomies of Mmu16 that can be used to identify the genetic basis of DS phenotypes where such genes reside in the region of Mmu16 orthologous to Hsa21. We use this panel to investigate the cardiac defects in DS, employing high-resolution episcopic microscopy (HREM) to analyze the precise three-dimensional (3D) morphology of the developing hearts. This combined approach enabled us to narrow down the critical region for CHD in DS to a minimal region containing 39 protein-coding genes and to show that this region contains at least two dosage-sensitive loci required in three copies to cause CHD. Furthermore using dual-wavelength HREM we show that the DMP develops normally in these DS models, and conclude that CHD in these DS models are not caused by failure in formation or growth of the DMP.

Cardiac abnormalities are very common in DS. Approximately half of all babies born with DS have a heart defect, many of which are serious and need to be surgically repaired. In order to understand the genetic and molecular mechanisms that lead to CHD in DS it is essential to establish mouse models for DS that accurately recapitulate them. Here we report the generation of a new mouse strain Dp1Tyb with a 23 Mb duplication of the entire region of Mmu16 orthologous to Hsa21, and show that it recapitulates the main types of CHD seen in DS. The Dp(16)1Yey mouse strain carries a similar duplication to Dp1Tyb encompassing the same set of genes and was also reported to have heart defects during embryogenesis (Li et al., 2007). Interestingly, the cardiac phenotypes in Dp1Tyb are very similar to the ones we previously observed in the transchromosomic strain Tc1 (Dunlevy et al., 2010; O'Doherty et al., 2005), despite differences in the Hsa21 orthologous gene content in the two strains and the different species origin for the duplicated genes: mouse in Dp1Tyb mice, human in Tc1 mice. This phenotypic similarity supports the conclusion that the CHD seen in these strains are caused by the same pathological mechanisms as those in people with DS. As with CHD in humans with DS we observed incomplete penetrance of the phenotype in Dp1Tyb mice. In humans this has been ascribed to genetic modifiers (Li et al., 2012). However the Dp1Tyb mouse strain was analyzed on an inbred background, suggesting that incomplete penetrance may be caused by stochastic effects during embryonic cardiogenesis. While the recovery of E14.5 Dp1Tyb embryos was consistent with expected Mendelian numbers (not shown), the recovery of Dp1Tyb mice at weaning was reduced to 50% of the expected numbers (Table 1). Some of this loss could be due to the most severe CHDs, since AVSDs were found in ~25% of E14.5 Dp1Tyb embryos, but it is likely that there are other unknown causes for this perinatal lethality. The other strain to show AVSDs, Dp3Tyb, showed a 25% reduction in recovery of mice at weaning, similar to the observed frequency of AVSDs at E14.5, making this a likely cause of the reduced yield of mutant mice.

Together with the Dp1Tyb strain we generated a collection of 7 new mouse strains with segmental duplications ranging from 1.5 Mb to 23 Mb that provide a genetic resource for mapping dosage-sensitive genes required in 3 copies to cause DS phenotypes. We used this mapping panel to identify a 4.9 Mb genomic region (from Mir802 to Zbtb21) that when present in 3 copies is sufficient to cause CHD. Furthermore, from analysis of shorter duplications, we show that the phenotype is caused by at least two distinct dosage-sensitive loci. By combining this data with analysis of the Ts1Rhr strain, we have determined that, minimally, one of the two loci lies either in an interval at the centromeric end of the duplication in Dp3Tyb (from Mir802 to Setd4) or at the telomeric end (from Mx2 to Zbtb21) (Figure 6a). These two segments between them contain only 7 coding genes (Setd4, Mx2, Tmprss2, Ripk4, Prdm15, C2cd2 and Zbtb21) and one microRNA gene (Mir802), none of which had been previously implicated in causing CHD in DS. A previous study proposed that a 3.7 Mb genomic region of Mmu16 (Ifnar1 – Kcnj6) in Dp(16)4Yey was sufficient to cause CHD (Liu et al., 2013); this region overlaps the centromeric end of the duplicated region of Dp3Tyb by 1.9 Mb from Mir802 to Kcnj6, which may help to further narrow down the search for causative genes (Figure 6b). We note that while the Dp5Tyb and Ts1Rhr strains did not show a statistically significant increase in CHDs compared to their wild-type littermate controls, in both strains we detected 2 mutant embryos with AVSDs (out of 20 or 21 embryos analyzed respectively). Thus it is possible that these strains have a weak phenotype – a larger number of embryos from these strains would need to be analyzed to establish whether this is significant. If true, it would suggest that one or more of the causative genes may reside within the Dp5Tyb interval (which is entirely contained within the Ts1Rhr interval).

Figure 6

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In contrast, analysis of humans with partial trisomies of Hsa21 identified distinct regions that contribute to cardiac abnormalities (Barlow et al., 2001; Korbel et al., 2009). An individual (PM) trisomic for the 7.7 Mb region from PSMG1 to PRMT2 had a VSD, whereas the shortest partial trisomy (individual BA) that gave rise to AVSD extended 10.0Mb from HLCS to PRMT2 (Figure 6b). We note that that while both of these intervals overlap substantially with the minimal Dp3Tyb region identified in our studies, the duplicated region in individual PM does not overlap at all with the Ifnar1 – Kcnj6 region identified by Liu et al as being sufficient to cause CHDs (Liu et al., 2013). In particular the human studies suggested that increased dosage of the DSCAM gene may be responsible for the CHDs (Korbel et al., 2009). In contrast the mouse studies show that duplication of Dscam is neither necessary or sufficient to cause CHDs, since the gene is not within the minimal Ifnar1 – Kcnj6 region identified by Liu et al as being sufficient for CHDs (Liu et al., 2013) but is duplicated in the Dp6Tyb strain which we show here does not have CHDs. Taking the human and mouse data together, this suggests that several dosage-sensitive genes may contribute to CHDs, with potentially no single gene being absolutely required.

Dp1Tyb mice show AVSD with ventricular but not atrial shunting. Analysis of AVSD in people with DS showed that the most common subtype was a complete AVSD with both atrial and ventricular shunting and the next most common was AVSD with exclusive ventricular shunting similar to that seen in the Dp1Tyb mouse (Freeman et al., 2008). Thus, although Dp1Tyb mice show AVSD, they only model one subtype of AVSD seen in people with DS – AVSD with ventricular shunting. It is not known why some cases of DS have complete AVSD whereas others have subtypes with only atrial or ventricular shunting, but it is tempting to speculate that there are distinct mechanisms driving defects in the atrial and ventricular septa.

AVSD have been shown to result from defects in development of the DMP (Briggs et al., 2013; Goddeeris et al., 2008; Rana et al., 2014; Tian et al., 2010; Webb et al., 1999; Xie et al., 2012). Importantly, a variety of different genetic manipulations that affect the DMP all resulted in AVSD with exclusive atrial shunting, demonstrating the importance of the DMP for the correct formation of the atrial side of the AV junction (Briggs et al., 2012). In contrast, in Dp1Tyb mice we found normal DMP development and no AVSD with atrial shunting, but instead saw exclusively AVSD with ventricular shunting (Figure 6c). Thus we propose that defects in the atrial septum causing the ostium primum type of AVSD are caused by malformation of the DMP, whereas ventricular defects are due to perturbations in other tissues, such as endocardial cushions or myocardium. Interestingly, Gata4-deficient mice display AVSD and have a normal DMP, but show perturbations in myocardial signaling to the adjacent atrioventricular cushion mesenchyme, suggesting that defects in myocardial signaling could lead to AVSD (Misra et al., 2014; Rajagopal et al., 2007). To our knowledge the Dp1Tyb and Dp3Tyb mouse strains are the first to have been shown to have AVSD with ventricular shunting and thus will allow direct studies of the pathological mechanisms underlying this subtype of AVSD.

In conclusion, we have generated a mapping panel of 7 mouse strains that can be used to identify dosage-sensitive genes underlying the broad range of DS phenotypes, and we have exploited this panel to map the location of genes causing CHD in DS.

1. 1. Adams DJ
   2. Biggs PJ
   3. Cox T
   4. Davies R
   5. van der Weyden L
   6. Jonkers J
   7. Smith J
   8. Plumb B
   9. Taylor R
   10. Nishijima I
   11. Yu Y
   12. Rogers J
   13. Bradley A

   (2004) Mutagenic insertion and chromosome engineering resource (mICER)

   Nature Genetics 36:867–871.

   https://doi.org/10.1038/ng1388

   • Google Scholar
2. 1. Anderson RH
   2. Mohun TJ
   3. Brown NA

   (2015) Clarifying the morphology of the ostium primum defect

   Journal of Anatomy 226:244–257.

   https://doi.org/10.1111/joa.12272

   • Google Scholar
3. 1. Baek KH
   2. Zaslavsky A
   3. Lynch RC
   4. Britt C
   5. Okada Y
   6. Siarey RJ
   7. Lensch MW
   8. Park IH
   9. Yoon SS
   10. Minami T
   11. Korenberg JR
   12. Folkman J
   13. Daley GQ
   14. Aird WC
   15. Galdzicki Z
   16. Ryeom S

   (2009) Down's syndrome suppression of tumour growth and the role of the calcineurin inhibitor DSCR1

   Nature 459:1126–1130.

   https://doi.org/10.1038/nature08062

   • Google Scholar
4. 1. Barlow GM
   2. Chen X-N
   3. Shi ZY
   4. Lyons GE
   5. Kurnit DM
   6. Celle L
   7. Spinner NB
   8. Zackai E
   9. Pettenati MJ
   10. Van Riper AJ
   11. Vekemans MJ
   12. Mjaatvedt CH
   13. Korenberg JR

   (2001) Down syndrome congenital heart disease: a narrowed region and a candidate gene

   Genetics in Medicine 3:91–101.

   https://doi.org/10.1097/00125817-200103000-00002

   • Google Scholar
5. 1. Blom NA
   2. Ottenkamp J
   3. Wenink AGC
   4. Gittenberger-de Groot AC

   (2003) Deficiency of the vestibular spine in atrioventricular septal defects in human fetuses with down syndrome

   The American Journal of Cardiology 91:180–184.

   https://doi.org/10.1016/S0002-9149(02)03106-5

   • Google Scholar
6. 1. Brault V
   2. Duchon A
   3. Romestaing C
   4. Sahun I
   5. Pothion S
   6. Karout M
   7. Borel C
   8. Dembele D
   9. Bizot JC
   10. Messaddeq N
   11. Sharp AJ
   12. Roussel D
   13. Antonarakis SE
   14. Dierssen M
   15. Hérault Y

   (2015) Opposite phenotypes of muscle strength and locomotor function in mouse models of partial trisomy and monosomy 21 for the proximal Hspa13-app region

   PLoS Genetics 11:e1005062.

   https://doi.org/10.1371/journal.pgen.1005062

   • Google Scholar
7. 1. Briggs LE
   2. Kakarla J
   3. Wessels A

   (2012) The pathogenesis of atrial and atrioventricular septal defects with special emphasis on the role of the dorsal mesenchymal protrusion

   Differentiation 84:117–130.

   https://doi.org/10.1016/j.diff.2012.05.006

   • Google Scholar
8. 1. Briggs LE
   2. Phelps AL
   3. Brown E
   4. Kakarla J
   5. Anderson RH
   6. van den Hoff MJB
   7. Wessels A

   (2013) Expression of the BMP receptor Alk3 in the second heart field is essential for development of the dorsal mesenchymal protrusion and atrioventricular septation

   Circulation Research 112:1420–1432.

   https://doi.org/10.1161/CIRCRESAHA.112.300821

   • Google Scholar
9. 1. Cai C-L
   2. Martin JC
   3. Sun Y
   4. Cui L
   5. Wang L
   6. Ouyang K
   7. Yang L
   8. Bu L
   9. Liang X
   10. Zhang X
   11. Stallcup WB
   12. Denton CP
   13. McCulloch A
   14. Chen J
   15. Evans SM
   16. Cui L
   17. Wang L
   18. Ouyang K
   19. Yang L
   20. Bu L
   21. Liang X
   22. Zhang X

   (2008) A myocardial lineage derives from Tbx18 epicardial cells

   Nature 454:104–108.

   https://doi.org/10.1038/nature06969

   • Google Scholar
10. 1. Chakrabarti L
    2. Best TK
    3. Cramer NP
    4. Carney RSE
    5. Isaac JTR
    6. Galdzicki Z
    7. Haydar TF

    (2010) Olig1 and Olig2 triplication causes developmental brain defects in down syndrome

    Nature Neuroscience 13:927–934.

    https://doi.org/10.1038/nn.2600

    • Google Scholar
11. 1. Davisson MT
    2. Schmidt C
    3. Akeson EC

    (1990)

    Segmental trisomy of murine chromosome 16: a new model system for studying down syndrome

    Progress in Clinical and Biological Research 360:263–280.

    • Google Scholar
12. 1. Delabar JM
    2. Theophile D
    3. Rahmani Z
    4. Chettouh Z
    5. Blouin JL
    6. Prieur M
    7. Noel B
    8. Sinet PM

    (1993)

    Molecular mapping of twenty-four features of down syndrome on chromosome 21

    European Journal of Human Genetics 1:114–124.

    • Google Scholar
13. 1. Duchon A
    2. Raveau M
    3. Chevalier C
    4. Nalesso V
    5. Sharp AJ
    6. Herault Y

    (2011) Identification of the translocation breakpoints in the Ts65Dn and Ts1Cje mouse lines: relevance for modeling down syndrome

    Mammalian Genome 22:674–684.

    https://doi.org/10.1007/s00335-011-9356-0

    • Google Scholar
14. 1. Dunlevy L
    2. Bennett M
    3. Slender A
    4. Lana-Elola E
    5. Tybulewicz VL
    6. Fisher EMC
    7. Mohun T

    (2010) Down's syndrome-like cardiac developmental defects in embryos of the transchromosomic Tc1 mouse

    Cardiovascular Research 88:287–295.

    https://doi.org/10.1093/cvr/cvq193

    • Google Scholar
15. 1. Fahed AC
    2. Gelb BD
    3. Seidman JG
    4. Seidman CE

    (2013) Genetics of congenital heart disease: the glass half empty

    Circulation Research 112:707–720.

    https://doi.org/10.1161/CIRCRESAHA.112.300853

    • Google Scholar
16. 1. Freeman SB
    2. Taft LF
    3. Dooley KJ
    4. Allran K
    5. Sherman SL
    6. Hassold TJ
    7. Khoury MJ
    8. Saker DM

    (1998) Population-based study of congenital heart defects in down syndrome

    American Journal of Medical Genetics 80:213–217.

    https://doi.org/10.1002/(SICI)1096-8628(19981116)80:3<213::AID-AJMG6>3.0.CO;2-8

    • Google Scholar
17. 1. Freeman SB
    2. Bean LH
    3. Allen EG
    4. Tinker SW
    5. Locke AE
    6. Druschel C
    7. Hobbs CA
    8. Romitti PA
    9. Royle MH
    10. Torfs CP
    11. Dooley KJ
    12. Sherman SL

    (2008) Ethnicity, sex, and the incidence of congenital heart defects: a report from the national down syndrome project

    Genetics in Medicine 10:173–180.

    https://doi.org/10.1097/GIM.0b013e3181634867

    • Google Scholar
18. 1. Goddeeris MM
    2. Rho S
    3. Petiet A
    4. Davenport CL
    5. Johnson GA
    6. Meyers EN
    7. Klingensmith J

    (2008) Intracardiac septation requires hedgehog-dependent cellular contributions from outside the heart

    Development 135:1887–1895.

    https://doi.org/10.1242/dev.016147

    • Google Scholar
19. 1. Gribble SM
    2. Wiseman FK
    3. Clayton S
    4. Prigmore E
    5. Langley E
    6. Yang F
    7. Maguire S
    8. Fu B
    9. Rajan D
    10. Sheppard O
    11. Scott C
    12. Hauser H
    13. Stephens PJ
    14. Stebbings LA
    15. Ng BL
    16. Fitzgerald T
    17. Quail MA
    18. Banerjee R
    19. Rothkamm K
    20. Tybulewicz VLJ
    21. Fisher EMC
    22. Carter NP
    23. Thomas T
    24. Maguire S
    25. Fu B
    26. Rajan D
    27. Sheppard O

    (2013) Massively parallel sequencing reveals the complex structure of an irradiated human chromosome on a mouse background in the Tc1 model of down syndrome

    PLoS ONE 8:e60482.

    https://doi.org/10.1371/journal.pone.0060482

    • Google Scholar
20. Book

    1. His W

    (1880)

    Die area interposita, die Eustachische klappe und die spina vestibuli

    In: His W, editors. Anatomie Menschlicher Embryonen III Zur Geschichte Der Organe. Leipzig, Germany: Verlag von FCWVogel. pp. 149–152.

    • Google Scholar
21. 1. Korbel JO
    2. Tirosh-Wagner T
    3. Urban AE
    4. Chen X-N
    5. Kasowski M
    6. Dai L
    7. Grubert F
    8. Erdman C
    9. Gao MC
    10. Lange K
    11. Sobel EM
    12. Barlow GM
    13. Aylsworth AS
    14. Carpenter NJ
    15. Clark RD
    16. Cohen MY
    17. Doran E
    18. Falik-Zaccai T
    19. Lewin SO
    20. Lott IT
    21. McGillivray BC
    22. Moeschler JB
    23. Pettenati MJ
    24. Pueschel SM
    25. Rao KW
    26. Shaffer LG
    27. Shohat M
    28. Van Riper AJ
    29. Warburton D
    30. Weissman S
    31. Gerstein MB
    32. Snyder M
    33. Korenberg JR
    34. Lange K

    (2009) The genetic architecture of down syndrome phenotypes revealed by high-resolution analysis of human segmental trisomies

    Proceedings of the National Academy of Sciences of the United States of America 106:12031–12036.

    https://doi.org/10.1073/pnas.0813248106

    • Google Scholar
22. 1. Korenberg JR
    2. Chen XN
    3. Schipper R
    4. Sun Z
    5. Gonsky R
    6. Gerwehr S
    7. Carpenter N
    8. Daumer C
    9. Dignan P
    10. Disteche C

    (1994) Down syndrome phenotypes: the consequences of chromosomal imbalance

    Proceedings of the National Academy of Sciences of the United States of America 91:4997–5001.

    https://doi.org/10.1073/pnas.91.11.4997

    • Google Scholar
23. 1. Lana-Elola E
    2. Watson-Scales SD
    3. Fisher EMC
    4. Tybulewicz VLJ

    (2011) Down syndrome: searching for the genetic culprits

    Disease Models & Mechanisms 4:586–595.

    https://doi.org/10.1242/dmm.008078

    • Google Scholar
24. 1. Li Z
    2. Yu T
    3. Morishima M
    4. Pao A
    5. LaDuca J
    6. Conroy J
    7. Nowak N
    8. Matsui S-I
    9. Shiraishi I
    10. Yu YE

    (2007) Duplication of the entire 22.9 mb human chromosome 21 syntenic region on mouse chromosome 16 causes cardiovascular and gastrointestinal abnormalities

    Human Molecular Genetics 16:1359–1366.

    https://doi.org/10.1093/hmg/ddm086

    • Google Scholar
25. 1. Li H
    2. Cherry S
    3. Klinedinst D
    4. DeLeon V
    5. Redig J
    6. Reshey B
    7. Chin MT
    8. Sherman SL
    9. Maslen CL
    10. Reeves RH

    (2012) Genetic modifiers predisposing to congenital heart disease in the sensitized down syndrome population

    Circulation: Cardiovascular Genetics 5:301–308.

    https://doi.org/10.1161/CIRCGENETICS.111.960872

    • Google Scholar
26. 1. Liu C
    2. Morishima M
    3. Yu T
    4. Matsui S
    5. Zhang L
    6. Fu D
    7. Pao A
    8. Costa AC
    9. Gardiner KJ
    10. Cowell JK
    11. Nowak NJ
    12. Nowak NJ
    13. Parmacek MS
    14. Liang P
    15. Baldini A
    16. Yu YE
    17. Matsui SI
    18. Zhang L
    19. Fu D
    20. Pao A
    21. Costa AC
    22. Gardiner KJ
    23. Cowell JK

    (2011) Genetic analysis of down syndrome-associated heart defects in mice

    Human Genetics 130:623–632.

    https://doi.org/10.1007/s00439-011-0980-2

    • Google Scholar
27. 1. Liu C
    2. Morishima M
    3. Jiang X
    4. Yu T
    5. Meng K
    6. Ray D
    7. Pao A
    8. Ye P
    9. Parmacek MS
    10. Yu YE

    (2013) Engineered chromosome-based genetic mapping establishes a 3.7 mb critical genomic region for down syndrome-associated heart defects in mice

    Human Genetics 133:743–753.

    https://doi.org/10.1007/s00439-013-1407-z

    • Google Scholar
28. 1. Lyle R
    2. Béna F
    3. Gagos S
    4. Gehrig C
    5. Lopez G
    6. Schinzel A
    7. Lespinasse J
    8. Bottani A
    9. Dahoun S
    10. Taine L
    11. Doco-Fenzy M
    12. Cornillet-Lefèbvre P
    13. Pelet A
    14. Lyonnet S
    15. Toutain A
    16. Colleaux L
    17. Horst J
    18. Kennerknecht I
    19. Wakamatsu N
    20. Descartes M
    21. Franklin JC
    22. Florentin-Arar L
    23. Kitsiou S
    24. Aït Yahya-Graison E
    25. Costantine M
    26. Sinet P-M
    27. Delabar JM
    28. Antonarakis SE

    (2009) Genotype–phenotype correlations in down syndrome identified by array CGH in 30 cases of partial trisomy and partial monosomy chromosome 21

    European Journal of Human Genetics 17:454–466.

    https://doi.org/10.1038/ejhg.2008.214

    • Google Scholar
29. 1. Magin TM
    2. McWhir J
    3. Melton DW

    (1992) A new mouse embryonic stem cell line with good germ line contribution and gene targeting frequency

    Nucleic Acids Research 20:3795–3796.

    https://doi.org/10.1093/nar/20.14.3795

    • Google Scholar
30. 1. Mahle WT
    2. Shirali GS
    3. Anderson RH

    (2006) Echo-morphological correlates in patients with atrioventricular septal defect and common atrioventricular junction

    Cardiology in the Young 16:43–51.

    https://doi.org/10.1017/S1047951106000758

    • Google Scholar
31. 1. Misra C
    2. Chang S-W
    3. Basu M
    4. Huang N
    5. Garg V

    (2014) Disruption of myocardial Gata4 and Tbx5 results in defects in cardiomyocyte proliferation and atrioventricular septation

    Human Molecular Genetics 23:5025–5035.

    https://doi.org/10.1093/hmg/ddu215

    • Google Scholar
32. 1. Mohun TJ
    2. Weninger WJ

    (2011) Imaging heart development using high-resolution episcopic microscopy

    Current Opinion in Genetics & Development 21:573–578.

    https://doi.org/10.1016/j.gde.2011.07.004

    • Google Scholar
33. 1. Mommersteeg MT
    2. Soufan AT
    3. de Lange FJ
    4. van den Hoff MJ
    5. Anderson RH
    6. Christoffels VM
    7. Moorman AF

    (2006) Two distinct pools of mesenchyme contribute to the development of the atrial septum

    Circulation Research 99:351–353.

    https://doi.org/10.1161/01.RES.0000238360.33284.a0

    • Google Scholar
34. 1. O'Doherty A
    2. Ruf S
    3. Mulligan C
    4. Hildreth V
    5. Errington ML
    6. Cooke S
    7. Sesay A
    8. Modino S
    9. Vanes L
    10. Hernandez D
    11. Linehan JM
    12. Sharpe PT
    13. Brandner S
    14. Bliss TV
    15. Henderson DJ
    16. Nizetic D
    17. Tybulewicz VL
    18. Fisher EM

    (2005) An aneuploid mouse strain carrying human chromosome 21 with down syndrome phenotypes

    Science 309:2033–2037.

    https://doi.org/10.1126/science.1114535

    • Google Scholar
35. 1. Olson LE
    2. Richtsmeier JT
    3. Leszl J
    4. Reeves RH

    (2004) A chromosome 21 critical region does not cause specific down syndrome phenotypes

    Science 306:687–690.

    https://doi.org/10.1126/science.1098992

    • Google Scholar
36. 1. Pereira PL
    2. Magnol L
    3. Sahun I
    4. Brault V
    5. Duchon A
    6. Prandini P
    7. Gruart A
    8. Bizot J-C
    9. Chadefaux-Vekemans B
    10. Deutsch S
    11. Trovero F
    12. Delgado-Garcia JM
    13. Antonarakis SE
    14. Dierssen M
    15. Herault Y
    16. Duchon A
    17. Prandini P
    18. Gruart A
    19. Bizot JC
    20. Chadefaux-Vekemans B
    21. Deutsch S

    (2009) A new mouse model for the trisomy of the Abcg1-U2af1 region reveals the complexity of the combinatorial genetic code of down syndrome

    Human Molecular Genetics 18:4756–4769.

    https://doi.org/10.1093/hmg/ddp438

    • Google Scholar
37. 1. Rajagopal SK
    2. Ma Q
    3. Obler D
    4. Shen J
    5. Manichaikul A
    6. Tomita-Mitchell A
    7. Boardman K
    8. Briggs C
    9. Garg V
    10. Srivastava D
    11. Goldmuntz E
    12. Broman KW
    13. Woodrow Benson D
    14. Smoot LB
    15. Pu WT
    16. Briggs C
    17. Garg V
    18. Srivastava D

    (2007) Spectrum of heart disease associated with murine and human GATA4 mutation

    Journal of Molecular and Cellular Cardiology 43:677–685.

    https://doi.org/10.1016/j.yjmcc.2007.06.004

    • Google Scholar
38. 1. Rana MS
    2. Theveniau-Ruissy M
    3. De Bono C
    4. Mesbah K
    5. Francou A
    6. Rammah M
    7. Dominguez JN
    8. Roux M
    9. Laforest B
    10. Anderson RH
    11. Mohun T
    12. Zaffran S
    13. Christoffels VM
    14. Kelly RG

    (2014) Tbx1 coordinates addition of posterior second heart field progenitor cells to the arterial and venous poles of the heart

    Circulation Research 115:790–799.

    https://doi.org/10.1161/CIRCRESAHA.115.305020

    • Google Scholar
39. 1. Rosset A
    2. Spadola L
    3. Ratib O

    (2004) OsiriX: an open-source software for navigating in multidimensional DICOM images

    Journal of Digital Imaging 17:205–216.

    https://doi.org/10.1007/s10278-004-1014-6

    • Google Scholar
40. 1. Sago H
    2. Carlson EJ
    3. Smith DJ
    4. Kilbridge J
    5. Rubin EM
    6. Mobley WC
    7. Epstein CJ
    8. Huang T-T

    (1998) Ts1Cje, a partial trisomy 16 mouse model for down syndrome, exhibits learning and behavioral abnormalities

    Proceedings of the National Academy of Sciences of the United States of America 95:6256–6261.

    https://doi.org/10.1073/pnas.95.11.6256

    • Google Scholar
41. 1. Salehi A
    2. Delcroix J-D
    3. Belichenko PV
    4. Zhan K
    5. Wu C
    6. Valletta JS
    7. Takimoto-Kimura R
    8. Kleschevnikov AM
    9. Sambamurti K
    10. Chung PP
    11. Xia W
    12. Villar A
    13. Campbell WA
    14. Kulnane LS
    15. Nixon RA
    16. Lamb BT
    17. Epstein CJ
    18. Stokin GB
    19. Goldstein LSB
    20. Mobley WC
    21. Valletta JS
    22. Takimoto-Kimura R
    23. Kleschevnikov AM
    24. Sambamurti K
    25. Chung PP

    (2006) Increased app expression in a mouse model of down's syndrome disrupts NGF transport and causes cholinergic neuron degeneration

    Neuron 51:29–42.

    https://doi.org/10.1016/j.neuron.2006.05.022

    • Google Scholar
42. 1. Snarr BS
    2. O'Neal JL
    3. Chintalapudi MR
    4. Wirrig EE
    5. Phelps AL
    6. Kubalak SW
    7. Wessels A

    (2007a) Isl1 expression at the venous pole identifies a novel role for the second heart field in cardiac development

    Circulation Research 101:971–974.

    https://doi.org/10.1161/CIRCRESAHA.107.162206

    • Google Scholar
43. 1. Snarr BS
    2. Wirrig EE
    3. Phelps AL
    4. Trusk TC
    5. Wessels A

    (2007) A spatiotemporal evaluation of the contribution of the dorsal mesenchymal protrusion to cardiac development

    Developmental Dynamics 236:1287–1294.

    https://doi.org/10.1002/dvdy.21074

    • Google Scholar
44. 1. Soriano P

    (1999) Generalized lacZ expression with the ROSA26 cre reporter strain

    Nature Genetics 21:70–71.

    https://doi.org/10.1038/5007

    • Google Scholar
45. 1. Sun Y
    2. Liang X
    3. Najafi N
    4. Cass M
    5. Lin L
    6. Cai C-L
    7. Chen J
    8. Evans SM

    (2007) Islet 1 is expressed in distinct cardiovascular lineages, including pacemaker and coronary vascular cells

    Developmental Biology 304:286–296.

    https://doi.org/10.1016/j.ydbio.2006.12.048

    • Google Scholar
46. 1. Sussan TE
    2. Yang A
    3. Li F
    4. Ostrowski MC
    5. Reeves RH

    (2008) Trisomy represses ApcMin-mediated tumours in mouse models of down’s syndrome

    Nature 451:73–75.

    https://doi.org/10.1038/nature06446

    • Google Scholar
47. 1. Tang S-HE
    2. Silva FJ
    3. Tsark WMK
    4. Mann JR

    (2002) A cre/loxP-deleter transgenic line in mouse strain 129S1/SvImJ

    Genesis 32:199–202.

    https://doi.org/10.1002/gene.10030

    • Google Scholar
48. 1. Tian Y
    2. Yuan L
    3. Goss AM
    4. Wang T
    5. Yang J
    6. Lepore JJ
    7. Zhou D
    8. Schwartz RJ
    9. Patel V
    10. Cohen ED
    11. Morrisey EE

    (2010) Characterization and in vivo pharmacological rescue of a Wnt2-Gata6 pathway required for cardiac inflow tract development

    Developmental Cell 18:275–287.

    https://doi.org/10.1016/j.devcel.2010.01.008

    • Google Scholar
49. 1. Vis JC
    2. Duffels MGJ
    3. Winter MM
    4. Weijerman ME
    5. Cobben JM
    6. Huisman SA
    7. Mulder BJM

    (2009) Down syndrome: a cardiovascular perspective

    Journal of Intellectual Disability Research 53:419–425.

    https://doi.org/10.1111/j.1365-2788.2009.01158.x

    • Google Scholar
50. 1. Webb S
    2. Brown NA
    3. Anderson RH

    (1998) Formation of the atrioventricular septal structures in the normal mouse

    Circulation Research 82:645–656.

    https://doi.org/10.1161/01.RES.82.6.645

    • Google Scholar
51. 1. Webb S
    2. Anderson RH
    3. Lamers WH
    4. Brown NA

    (1999) Mechanisms of deficient cardiac septation in the mouse with trisomy 16

    Circulation Research 84:897–905.

    https://doi.org/10.1161/01.RES.84.8.897

    • Google Scholar
52. 1. Weninger WJ
    2. Geyer SH
    3. Mohun TJ
    4. Rasskin-Gutman D
    5. Matsui T
    6. Ribeiro I
    7. Costa Luciano da F.
    8. Izpisúa-Belmonte JC
    9. Müller GB

    (2006) High-resolution episcopic microscopy: a rapid technique for high detailed 3D analysis of gene activity in the context of tissue architecture and morphology

    Anatomy and Embryology 211:213–221.

    https://doi.org/10.1007/s00429-005-0073-x

    • Google Scholar
53. 1. Xie L
    2. Hoffmann AD
    3. Burnicka-Turek O
    4. Friedland-Little JM
    5. Zhang K
    6. Moskowitz IP

    (2012) Tbx5-hedgehog molecular networks are essential in the second heart field for atrial septation

    Developmental Cell 23:280–291.

    https://doi.org/10.1016/j.devcel.2012.06.006

    • Google Scholar
54. 1. Yu Y
    2. Bradley A

    (2001) Mouse genomic technologies: engineering chromosomal rearrangements in mice

    Nature Reviews Genetics 2:780–790.

    https://doi.org/10.1038/35093564

    • Google Scholar
55. 1. Yu T
    2. Li Z
    3. Jia Z
    4. Clapcote SJ
    5. Liu C
    6. Li S
    7. Asrar S
    8. Pao A
    9. Chen R
    10. Fan N
    11. Carattini-Rivera S
    12. Bechard AR
    13. Spring S
    14. Henkelman RM
    15. Stoica G
    16. Matsui S-I
    17. Nowak NJ
    18. Roder JC
    19. Chen C
    20. Bradley A
    21. Yu YE
    22. Jia Z
    23. Clapcote SJ
    24. Liu C
    25. Li S
    26. Asrar S
    27. Pao A
    28. Chen R
    29. Fan N

    (2010) A mouse model of down syndrome trisomic for all human chromosome 21 syntenic regions

    Human Molecular Genetics 19:2780–2791.

    https://doi.org/10.1093/hmg/ddq179

    • Google Scholar

Author details

1. Eva Lana-Elola

   The Francis Crick Institute, London, United Kingdom

   Contribution

   ELE, Conception and design, Acquisition of data, Analysis and interpretation of data, Drafting or revising the article

   Competing interests

   The authors declare that no competing interests exist.

2. Sheona Watson-Scales

   The Francis Crick Institute, London, United Kingdom

   Contribution

   SWS, Conception and design, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

3. Amy Slender

   The Francis Crick Institute, London, United Kingdom

   Contribution

   AS, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

4. Dorota Gibbins

   The Francis Crick Institute, London, United Kingdom

   Contribution

   DG, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

5. Alexandrine Martineau

   The Francis Crick Institute, London, United Kingdom

   Contribution

   AM, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

6. Charlotte Douglas

   The Francis Crick Institute, London, United Kingdom

   Contribution

   CD, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

7. Timothy Mohun

   The Francis Crick Institute, London, United Kingdom

   Contribution

   TM, Conception and design, Analysis and interpretation of data, Drafting or revising the article

   Competing interests

   The authors declare that no competing interests exist.

8. Elizabeth MC Fisher

   Department of Neurodegenerative Disease, UCL Institute of Neurology, London, United Kingdom

   Contribution

   EMCF, Conception and design, Drafting or revising the article

   Competing interests

   The authors declare that no competing interests exist.

9. Victor LJ Tybulewicz

   1. The Francis Crick Institute, London, United Kingdom
   2. Imperial College London, London, United Kingdom

   Contribution

   VLJT, Conception and design, Drafting or revising the article

   For correspondence

   Victor.T@crick.ac.uk

   Competing interests

   The authors declare that no competing interests exist.

   "This ORCID iD identifies the author of this article:" 0000-0003-2439-0798

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