Disruption of DDX53 coding sequence has limited impact on iPSC-derived human NGN2 neurons.
BMC Med Genomics
; 16(1): 5, 2023 01 12.
Article
em En
| MEDLINE
| ID: mdl-36635662
ABSTRACT
BACKGROUND:
The X-linked PTCHD1 locus is strongly associated with autism spectrum disorder (ASD). Males who carry chromosome microdeletions of PTCHD1 antisense long non-coding RNA (PTCHD1-AS)/DEAD-box helicase 53 (DDX53) have ASD, or a sub-clinical form called Broader Autism Phenotype. If the deletion extends beyond PTCHD1-AS/DDX53 to the next gene, PTCHD1, which is protein-coding, the individuals typically have ASD and intellectual disability (ID). Three male siblings with a 90 kb deletion that affects only PTCHD1-AS (and not including DDX53) have ASD. We performed a functional analysis of DDX53 to examine its role in NGN2 neurons.METHODS:
We used the clustered regularly interspaced short palindromic repeats (CRISPR) gene editing strategy to knock out DDX53 protein by inserting 3 termination codons (3TCs) into two different induced pluripotent stem cell (iPSC) lines. DDX53 CRISPR-edited iPSCs were differentiated into cortical excitatory neurons by Neurogenin 2 (NGN-2) directed differentiation. The functional differences of DDX53-3TC neurons compared to isogenic control neurons with molecular and electrophysiological approaches were assessed.RESULTS:
Isogenic iPSC-derived control neurons exhibited low levels of DDX53 transcripts. Transcriptional analysis revealed the generation of excitatory cortical neurons and DDX53 protein was not detected in iPSC-derived control neurons by western blot. Control lines and DDX53-3TC neurons were active in the multi-electrode array, but no overt electrophysiological phenotype in either isogenic line was observed.CONCLUSION:
DDX53-3TC mutation does not alter NGN2 neuronal function in these experiments, suggesting that synaptic deficits causing ASD are unlikely in this cell type.Palavras-chave
Texto completo:
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Base de dados:
MEDLINE
Assunto principal:
RNA Helicases DEAD-box
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Células-Tronco Pluripotentes Induzidas
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Transtorno do Espectro Autista
Limite:
Humans
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Male
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article