RNA-seq of human induced pluripotent cells treated with short-term Met deprival

Human iPSCs require high amounts of methionine (Met). Met deprivation results in a rapid decrease in intracellular S-adenosyl-methionine (SAM), poising human iPSCs for differentiation and leading to apoptosis of undifferentiated cells. Met deprivation triggers a rapid metabolite change, including SAM, followed by reversible epigenetic modification. We show here that short-term Met deprivation impairs pluripotency network through epigenetic modification. The trimethylation of lysine 4 on histone H3 (H3K4me3) was dramatically affected compared to other histone modifications. Transcription start site (TSS) region of key pluripotent genes such as NANOG and OCT3/4 are specifically impacted upon short-term Met deprivation. Gene expression levels of these genes decreased, correlating to the loss of H3K4me3 marks. Upon differentiation, Met deprivation triggers a loss of the H3K27me3 in many mesendodermal genes, thereby switching from a bivalent to monovalent (H3K4me3) state. We concluded that Met metabolism maintains the pluripotent network with histone marks, and their loss potentiates differentiation. RNA-seq (n=3) in undifferentiated Ff-I01s01, an HLA homozygote donor-derived human iPSCs and the cells initiated a differentiation state with 100ng/mL Activin A and 3 µM CHIR-99021 for 24 hr. The cells formed spheres on a rotating orbital shaker for 24 hr, followed by a pre-cultured in complete or Met deprived medium for 5hr.