Journal: Advanced Science

Article Title: The PERK Branch of the Unfolded Protein Response Safeguards Protein Homeostasis and Mesendoderm Specification of Human Pluripotent Stem Cells

doi: 10.1002/advs.202303799

Figure Lengend Snippet: PERK regulates mesoderm and endoderm specification. A–C) Representative (left) and quantitative (right) immunofluorescence analysis of the mesoderm markers Brachyury and ISL1 A), definitive endoderm markers FOXA2 and SOX17 B), and ectoderm markers PAX6 and SOX2 C) in cultures derived from WT and PERK KO hESCs. n = 6 biologically independent experiments. Scale bars, 100 µm. ** P <0.01, *** P <0.001 versus WT. D–F) RT‐qPCR analysis of mesoderm D), definitive endoderm E), and ectoderm F) marker gene expression in cultures derived from WT and PERK KO hESCs. n = 6 biologically independent experiments. * P <0.05, *** P <0.001 versus WT. G) Hematoxylin and eosin staining of teratoma derived from WT and PERK KO hESCs, showing tissue derivatives of the mesoderm (left), definitive endoderm (middle), or ectoderm (right). Scale bars, 100 µm. Data represent mean ± SD. Statistical significance was determined by one‐way ANOVA with a post‐hoc Tukey test.

Article Snippet: Primary antibodies included SOX2 (1:200, Abcam, ab79351), NANOG (1:200, Cell Signaling Technology, 3 580), OCT4 (1:200, Santa Cruz, sc‐5279), Ki67 (1:100, BD, 550 609), cTNT (1:200, Thermo Fisher, MA5‐12960), α‐actinin (1:200, Sigma‐Aldrich, A773225), MEF2C (1:100, Cell Signaling Technology, D80C1), SOX17 (1:200, R&D, AF‐1924), FOXA2 (1:200, Cell Signaling Technology, 8186), PAX6 (1:100, Thermo Fisher, MA1‐109), ISL1 (1:50, DSHB), Brachyury (1:100, R&D, AF2085), Ubiquitin (P37) (1:200, Cell Signaling Technology, #58 395), ATF4 (1:200, Cell Signaling Technology, 11 815), and GRP78 (1:200, Abclonal, A23453).

Techniques: Immunofluorescence, Derivative Assay, Quantitative RT-PCR, Marker, Gene Expression, Staining