Selenoprotein T (SELENOT, SelT), a thioredoxin-like enzyme, exerts an essential oxidoreductase activity in the endoplasmic reticulum. However, its precise function remains unknown. To gain more understanding of SELENOT function, a conventional global Selenot knockout (KO) mouse model was constructed for the first time using the CRISPR/Cas9 technique. Deletion of SELENOT caused male sterility, reduced size/body weight, lower fed and/or fasting blood glucose levels and lower fasting serum insulin levels, and improved blood lipid profile. Tandem mass tag (TMT) proteomics analysis was conducted to explore the differentially expressed proteins (DEPs) in the liver of male mice, revealing 60 up-regulated and 94 down-regulated DEPs in KO mice. The proteomic results were validated by western blot of three selected DEPs. The elevated expression of Glycogen [starch] synthase, liver (Gys2) is consistent with the hypoglycemic phenotype in KO mice. Furthermore, the bioinformatics analysis showed that Selenot-KO-induced DEPs were mainly related to lipid metabolism, cancer, peroxisome proliferator-activated receptor (PPAR) signaling pathway, complement and coagulation cascades, and protein digestion and absorption. Overall, these findings provide a holistic perspective into SELENOT function and novel insights into the role of SELENOT in glucose and lipid metabolism, and thus, enhance our understanding of SELENOT function.

Background: Residual undifferentiated induced pluripotent stem cells (iPSCs) detection is essential for both Embryonic Stem Cells (ESCs) and iPSCs application in final cell therapy products. However, specific differentiated cells require specific genes for residual detection; identifying the suitable marker is costly and time-consuming. Thus, a universal marker for iPSCs residue detection for all three germline cells would greatly benefit PSC-derived cellular therapies. Methods: Next-generation sequencing (NGS) was performed on total RNAs isolated from the iPSC cell lines and embryonic stem cells (H9), the top 30 expressed genes were selected as candidates. By analysis expression fold change comparing iPSC cells to the differentiated cells, seven genes were highly expressed in iPSCs but showed minimal background expression in differentiated cells. Tissue expression pattern of the candidate genes were explored in the Genotype-Tissue Expression (GTEx) project database, candidate genes were narrowed down to two genes. Spike-in experiments were performed to determine the detection limit and correlation with the number of iPSCs and gene expression by ddPCR. Results: By next-generation sequencing (NGS), we identified two marker genes (ESRG and ZSCAN10) suitable for universal undifferentiated iPSC detection. Both ESRG and ZSCAN10 are highly expressed in iPSCs. ZSCAN10 is slightly expressed in the testis, pituitary, and cerebellum; ESRG is highly expressed in the vagina and scarcely expressed in the other tissues. Furthermore, the ddPCR method with a probe and primers for ESRG and ZSCAN10 detected a trace of undifferentiated hiPSCs to a spiked level of 0.0001%. Conclusions: These results suggest that targeting ESRG/ZSCAN10 transcripts is highly sensitive, quantitative, and could be broadly applied to quality control of almost all iPSC-derived cell therapy products.

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