In addition to showing that extracranial AVMs demonstrate interrupted elastin and that AVMs and LMs demonstrate abnormal α-smooth muscle actin just as brain AVMS do, our results demonstrate that NOTCH1, 2, 3 and 4 proteins are overexpressed to varying degrees in both the endothelial and mural lining of the malformed vessels in all types of malformations.
We identified novel co-segregating pathogenic mutations in NOTCH1 associated with left and right-sided cardiac malformations in three independent families with a total of 15 affected individuals.
In the TOF patients, we found four copy number gains affecting three genes, of which two are important regulators of heart development (NOTCH1, ISL1) and one is located in a region associated with cardiac malformations (PRODH at 22q11).
These findings suggest that patient-specific iPS cells may provide molecular insights into complex transcriptional and epigenetic mechanisms, at least in part, through combinatorial expression of NKX2-5, HAND1, and NOTCH1 that coordinately contribute to cardiac malformations in HLHS.
We recently identified missense variants in the NOTCH1 receptor in patients with diverse left ventricular outflow tract (LVOT) malformations (NOTCH1(G661S) and NOTCH1(A683T)) that reduce ligand-induced Notch signaling.
This finding was supported by the discovery of a NOTCH1 frameshift mutation in an unrelated family with similar aortic valve disease, suggesting that NOTCH1 haploinsufficiency was a genetic cause of aortic valve malformations and calcification.
These results suggest that NOTCH1 mutations cause an early developmental defect in the aortic valve and a later de-repression of calcium deposition that causes progressive aortic valve disease.
Genetically inactivating 1 allele of either Ednra or Bmp4 significantly reduced the penetrance of maxillary malformation in both Six1 <sup>-/-</sup> and Six1 <sup>-/-</sup> Six2 <sup>+/-</sup> embryos, indicating that Six1 and Six2 regulate both endothelin and bone morphogenetic protein-4 signaling pathways to pattern the facial structures.
In conclusion, we show that a de novo variant in SIX1 in a patient with sensorineural hearing loss leads to cochleovestibular malformations and abnormalities of the CVN, without any other abnormalities.
Previous studies indicated that transcriptional complex SIX1/EYA1 may contribute to SHF development, and SIX1/EYA1 knockout mice exhibited a series of conotruncal malformations.
Nevertheless, further studies are necessary to understand if altered SIX1 expression may play a role in human development of kidney and urinary tract congenital anomalies.
We demonstrate that murine mutation of both Six1 and Eya1 recapitulated most features of human del22q11 syndromes, including craniofacial, cardiac outflow tract, and aortic arch malformations.
Human SALL1 is a homologue of the Drosophila region-specific homeotic gene sal, and is also known as a causative gene for Townes-Brocks syndrome, which is characterized by multi-organ malformations.
These findings uncover an essential role for Six1 in establishing a functionally normal ureter and provide new insights into the molecular basis of urinary tract malformations in BOR patients.
The present, yet preliminary, observation that renal and temporal bone malformations are less frequent in SIX1-related disease suggests a slightly different clinical profile compared to EYA1-related disease.