The RAB4A gene is a member of the largest group in the Ras superfamily of small GTPases, which regulate membrane trafficking. The encoded protein is associated with early endosomes and is involved in sorting and recycling. In this study, we generated induced pluripotent stem cells (iPSC) from a healthy individual by electroporation of peripheral blood mononuclear cells. We generated a RAB4A homozygous knockout human iPSC line via CRISPR/Cas9 gene editing. The iPSCs-RAB4A−/− had a normal karyotype, expressed pluripotency markers, and maintained trilineage differentiation potential.
Activator protein 1 (AP-1) is involved in cell fate determination and function. To monitor the AP-1 activity, we cloned a AP-1 binding sites fragment into the upstream of minimal TATA-box promoter, then a luciferase-GFP reporter (LuciGFP) was designated to the downstream of the promoter. With CRISPR/Cas9, the AP-1-LuciGFP reporter was introduced into AAVS1 locus, a safe harbor for gene editing. Thus, this AP-1-LuciGFP reporter cell line could be subjected to monitor the AP-1 activity during the cell differentiation, cell fate transition and disease modeling.
We have successfully derived a hiPSC line from PBMCs obtained from a 41-year-old infertile female. The patient’s karyotype, as determined by Bionano OGM, revealed complex chromosomal rearrangements, including 46,XX,ins(1;15)(p13.3;q22.31q26.1),inv(2)(p22.1p16.3),t(2;14)(q34;q12). Specifically, the episomal plasmids encoding key reprogramming factors OCT4, sh-p53, SOX2, KLF4, L-MYC, and LIN28 were applied to generate the integration-free hiPSC line, which was designated as TONGJIi001-A. This line exhibits typical hiPSC morphology, expresses core pluripotency markers and presents the ability to differentiate into all three germ layers in vitro. Collectively, hiPSC TONGJIi001-A provides a valuable resource for investigating the mechanisms underlying chromosomal structural abnormalities associated with infertility.
1q21.1 deletion has been identified as a risk factor related to not only mental disorders such as schizophrenia, but also congenital heart defects. However, at human cellular and molecular levels, it is still not known how this variant affects brain and heart development and contributes to the onset of these diseases.
Here, we generated induced pluripotent stem cells (iPSCs) from a patient with 1q21.1 deletion. The iPSCs expressed stemness markers and exhibited the ability to differentiate into three germ layers in vitro. These iPSCs will be useful tools to understand the pathophysiology of mental disorders and heart defects in humans.
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