A Human Hereditary Cardiomyopathy Shares a Genetic Substrate With Bicuspid Aortic Valve
Authors: Siguero-Álvarez Marcos , Salguero-Jiménez Alejandro , Grego-Bessa Joaquim , de la Barrera Jorge , MacGrogan Donal , Prados Belén , Sánchez-Sáez Fernando , Piñeiro-Sabarís Rebeca , Felipe-Medina Natalia , Torroja Carlos , Gómez Manuel José , Sabater-Molina María , Escribá Rubén , Richaud-Patin Ivonne , Iglesias-García Olalla , Sbroggio Mauro , Callejas Sergio , O’Regan Declan P. , McGurk Kathryn A. , Dopazo Ana , Giovinazzo Giovanna , Ibañez Borja , Monserrat Lorenzo , Pérez-Pomares José María , Sánchez-Cabo Fátima , Pendas Alberto M. , Raya Angel , Gimeno-Blanes Juan R. , de la Pompa José Luis 
Published: November 2022

Background:
The complex genetics underlying human cardiac disease is evidenced by its heterogenous manifestation, multigenic basis, and sporadic occurrence. These features have hampered disease modeling and mechanistic understanding. Here, we show that 2 structural cardiac diseases, left ventricular noncompaction (LVNC) and bicuspid aortic valve, can be caused by a set of inherited heterozygous gene mutations affecting the NOTCH ligand regulator MIB1 (MINDBOMB1) and cosegregating genes.

Methods:

We used CRISPR-Cas9 gene editing to generate mice harboring a nonsense or a missense
MIB1
mutation that are both found in LVNC families. We also generated mice separately carrying these
MIB1
mutations plus 5 additional cosegregating variants in the
ASXL3
,
APCDD1
,
TMX3, CEP192
, and
BCL7A
genes identified in these LVNC families by whole exome sequencing. Histological, developmental, and functional analyses of these mouse models were carried out by echocardiography and cardiac magnetic resonance imaging, together with gene expression profiling by RNA sequencing of both selected engineered mouse models and human induced pluripotent stem cell–derived cardiomyocytes. Potential biochemical interactions were assayed in vitro by coimmunoprecipitation and Western blot.

Results:

Mice homozygous for the
MIB1
nonsense mutation did not survive, and the mutation caused LVNC only in heteroallelic combination with a conditional allele inactivated in the myocardium. The heterozygous
MIB1
missense allele leads to bicuspid aortic valve in a NOTCH-sensitized genetic background. These data suggest that development of LVNC is influenced by genetic modifiers present in affected families, whereas valve defects are highly sensitive to NOTCH haploinsufficiency. Whole exome sequencing of LVNC families revealed single-nucleotide gene variants of
ASXL3
,
APCDD1
,
TMX3, CEP192
, and
BCL7A
cosegregating with the
MIB1
mutations and LVNC. In experiments with mice harboring the orthologous variants on the corresponding
Mib1
backgrounds, triple heterozygous
Mib1 Apcdd1 Asxl3
mice showed LVNC, whereas quadruple heterozygous
Mib1 Cep192 Tmx3;Bcl7a
mice developed bicuspid aortic valve and other valve-associated defects. Biochemical analysis suggested interactions between CEP192, BCL7A, and NOTCH. Gene expression profiling of mutant mouse hearts and human induced pluripotent stem cell–derived cardiomyocytes revealed increased cardiomyocyte proliferation and defective morphological and metabolic maturation.

Conclusions:
These findings reveal a shared genetic substrate underlying LVNC and bicuspid aortic valve in which MIB1-NOTCH variants plays a crucial role in heterozygous combination with cosegregating genetic modifiers.