Inducible SMARCAL1 knockdown in iPSC reveals a link between replication stress and altered expression of master differentiation genes

Autor: Veronica Marabitti, Nicolò Morina, Annapaola Franchitto, Giusj Monia Pugliese, Federico Salaris, Alessandro Rosa, Valentina Palermo, Pietro Pichierri
Jazyk: angličtina
Rok vydání: 2019
Předmět:
0301 basic medicine
DNA Repair
Transcription
Genetic
ipsc
replication stress
DNA damage
Induced Pluripotent Stem Cells
Neuroscience (miscellaneous)
lcsh:Medicine
Medicine (miscellaneous)
Biology
General Biochemistry
Genetics and Molecular Biology
S Phase
03 medical and health sciences
0302 clinical medicine
Immunology and Microbiology (miscellaneous)
Stress
Physiological
lcsh:Pathology
medicine
Humans
Cell Lineage
Phosphorylation
Induced pluripotent stem cell
Gene
Gene knockdown
dna replication
lcsh:R
DNA Helicases
DNA replication
Cell Differentiation
medicine.disease
Phenotype
Osteochondrodysplasia
Cell biology
Replication Stress
SIOD
iPSC
030104 developmental biology
Gene Expression Regulation
siod
Dysplasia
Gene Knockdown Techniques
dna damage
030217 neurology & neurosurgery
Research Article
lcsh:RB1-214
Zdroj: Disease Models & Mechanisms, Vol 12, Iss 10 (2019)
Disease Models & Mechanisms
Popis: Schimke immuno-osseous dysplasia is an autosomal recessive genetic osteochondrodysplasia characterized by dysmorphism, spondyloepiphyseal dysplasia, nephrotic syndrome and frequently T cell immunodeficiency. Several hypotheses have been proposed to explain the pathophysiology of the disease; however, the mechanism by which SMARCAL1 mutations cause the syndrome is elusive. Here, we generated a conditional SMARCAL1 knockdown model in induced pluripotent stem cells (iPSCs) to mimic conditions associated with the severe form the disease. Using multiple cellular endpoints, we characterized this model for the presence of phenotypes linked to the replication caretaker role of SMARCAL1. Our data show that conditional knockdown of SMARCAL1 in human iPSCs induces replication-dependent and chronic accumulation of DNA damage triggering the DNA damage response. Furthermore, they indicate that accumulation of DNA damage and activation of the DNA damage response correlates with increased levels of R-loops and replication-transcription interference. Finally, we provide evidence that SMARCAL1-deficient iPSCs maintain active DNA damage response beyond differentiation, possibly contributing to the observed altered expression of a subset of germ layer-specific master genes. Confirming the relevance of SMARCAL1 loss for the observed phenotypes, they are prevented or rescued after re-expression of wild-type SMARCAL1 in our iPSC model. In conclusion, our conditional SMARCAL1 knockdown model in iPSCs may represent a powerful model when studying pathogenetic mechanisms of severe Schimke immuno-osseous dysplasia.
Summary: This study reports the generation of an iPSC model to investigate phenotypic changes associated with loss of SMARCAL1, as found in severe forms of the genetic disease Schimke immuno-osseous dysplasia.
Databáze: OpenAIRE
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