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. 2020 Oct 16;7(23):2002602. doi: 10.1002/advs.202002602

Figure 6.

Figure 6

PRMT5 interacts with and methylates the transcriptional pausing factor SPT5 to affect its binding to the Bscl2 TSS. a) ChIP‐sequencing results showing co‐occupancy of PRMT5, SPT5, and RNA polymerase II (POL2) at the same TSS region (boxed region) of Bscl2 genome (shown in the bottom panel). Note that PRMT5 only occupies the region in differentiated (D1) but not undifferentiated (D0) 3T3‐L1 cell. SPT5 ChIP‐seq is based on results from mouse embryonic stem cell (mES) and POL2 ChIP‐seq is based on results from eWAT.[ 27 , 28 ] b,c) ChIP‐qPCR analysis of b) PRMT5 and c) SPT5 binding to the boxed region shown in a in undifferentiated (D0) and differentiated 3T3‐L1 cells (D2). d) SPT5 ChIP‐qPCR results on Prmt5 overexpressing 3T3‐L1 cells and control cells. e) SPT5 ChIP‐qPCR of Bscl2 promoter in SVF cells from WT (Prmt5flox/flox) and Prmt5AKO mice after 6 day of differentiation in vitro. f) Lysates from undifferentiated (D0) and differentiated 3T3‐L1 cells (D6) were immunoprecipitated (IP) with SPT5 antibody and blotted with SPT5 and PRMT5 antibodies showing association with PRMT5 only in differentiated cells. g) Proximity ligation assay (PLA) using PRMT5 and SPT5 antibodies in WT undifferentiated (D0) and differentiated SVF preadipocytes (D4) showing PRMT5 interacts with SPT5 in nucleus only in differentiated cells. h) Lysates from eWAT of WT (Prmt5flox/flox) and Prmt5AKO mice were immunoprecipitated (IP) with SYM10 antibody and blotted with SPT5 antibody. (t‐test: **p < 0.01).