PML nuclear bodies form a regulatory hub via liquid-liquid phase separation for TRIM33 control of Lefty1/2 genes in mouse embryonic stem cells [RNA-seq]

TRIM33 is a chromatin reader required for nodal signaling during mesendoderm differentiation, although differences in its function between mouse embryonic stem cells (mESCs) and differentiated mesendoderm cells are unknown. Here, we found that TRIM33 co-condenses with PML nuclear bodies (NBs) via liquid-liquid phase separation specifically in mESCs to mediate nodal signaling-directed transcription of Lefty1/2. Our findings show that TRIM33 puncta formation depends on the presence of PML NBs. TurboID proximity labeling further revealed that PML NBs recruit distinct sets of client proteins in NaAsO2-treated, untreated mESCs, and differentiated cells. TRIM33 and PML co-regulate Lefty1/2 expression, while PML NBs directly associate with the Lefty1/2 loci and regulate a gene cluster that includes Lefty1/2 loci specifically in mESCs. Moreover, TRIM33 association with chromatin depends on PML NBs. Thus, PML NBs serve as a hub for transcriptional regulation of Lefty1/2 by TRIM33 and other pluripotency factors in mESCs. Overall design: ESCs were cultured on gelatin coated plates with 15% serum ESC culture medium. mESCs untreated/treated with sodium arsenite treated (200 µM), activin A (50 ng/ml) or SB431542 (10 µM) for 2h were harvested for RNA-seq