BS38 Long noncoding RNA STX18-AS1 regulates in vitro cardiomyocyte differentiation

Research output: Contribution to conferenceAbstract


Introduction The long noncoding RNA STX18-AS1 located at chromosome 4p16 harbours single nucleotide polymorphisms (SNPs) significantly associated with Atrial Septal Defect (ASD) in genome-wide association studies (GWAS). We previously showed that knockout (KO) of STX18-AS1 using CRISPR in human embryonic stem cell-derived cardiomyocytes (hESC-CMs) delayed the programming of transition from cardiac mesoderm to early cardiomyocytes in vitro, and was accompanied by cognate differences in key cardiac transcription factors. Here, we present data on STX18-AS1 rescue to confirm our hypotheses regarding its critical role in regulating heart lineage specifications.

Methods To rescue the effects of STX18-AS1 KO, we transfected STX18-AS1 KO cells using pcDNA3.1 overexpression plasmid (‘Rescue’). For comparison of the rescuing effects, ‘Rescue’ hESCs were subjected to in vitro CM differentiation together with WT, KO, and STX18-AS1 knockdown (KD, CRISPR cell pool following puromycine selection) cells with the same protocol. Phenotype changes and transcription of key cardiac markers were evaluated at D0, D2, D4, D6, D8, and D15 of CM differentiation.

Results The overall transcription levels of STX18-AS1 in multiple cell lines were KO < ‘Rescue’ < KD< WT. Compared to KO cells, with a delayed beating start at D10-D15, ‘Rescue’ cells started beating earlier at D8-D10 but slightly later than WT cells at D6-D8. Corresponding to the phenotype observations, transcriptional data showed that key cardiac transcriptional factors and marker genes, including NKX2–5, TBX5, GATA4, and TNNT2, were upregulated in ‘Rescue’ cells compared to KO cells during D6-D15 (cardiac mesoderm specification into cardiac progenitors and cardiomyocytes) but less than WT cells. For the same period, the transcription of NKX2–5, TBX5, and GATA4 in‘Rescue’ cells and KD cells were comparable in reference to WT cells.

Original languageEnglish
Publication statusPublished - Jun 2022


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