Fig 2: PDLIM1 inhibits NF-?B-mediated gene expression.(a) Cytoplasmic and nuclear extracts from NIH3T3 cells clones, stably expressing Myc-PDLIM1 (#13, #25) or empty vector (control) were immunoblotted with anti-PDLIM1. Upper bands correspond to stably expressed c-Myc-tagged PDLIM1 and lower bands are endogenous PDLIM1. The purity of the fractionations was confirmed by blotting with anti-HSP90 (cytoplasm) or anti-LSD1 (nucleus). Western blots are representative of at least three independent experiments. (b) Real-time RT-PCR analysis of IL-6 in control NIH3T3 cells and NIH3T3 cells expressing PDLIM1 (#13, #25) stimulated for 2 h with LPS. Data are means ± SD from at least three independent experiments. **P < 0.01. (c) Real-time RT-PCR analysis of LPS-inducible genes in control NIH3T3 cells and NIH3T3 cells expressing PDLIM1 (#13, #25), unstimulated or stimulated with LPS for 2 h. Data are means ± SD from at least three independent experiments. *P < 0.05; **P < 0.01.

Fig 3: Proposed mechanisms of gene regulation mediated by single nucleotide polymorphism rs163184, Sp3 and Lsd1/Kdm1a.

Fig 4: KDM1A knockdown highly up-regulated TIMP1. (A and B) Detection of MMP9 and MMP2 levels by PCR (A) and western blotting (B) after si-KDM1A or si-NC transfection in IHH-4 and TPC1 cells. (C) Gelatin zymography analysis of MMP9 activity in IHH-4 and TPC1 cells after KDM1A knockdown. (D and E) qRT-PCR analysis of TIMP1, TIMP2, TIMP3 and TIMP4 expression levels in IHH-4 (D) and TPC1 cells (E) after KDM1A knockdown. (F) Western blot analysis of the effect of KDM1A knockdown on TIMP1 protein levels in IHH-4 and TPC1 cells. GAPDH was used as an internal loading control. (G) Correlation analysis between KDM1A and TIMP1 expressions in 60 papillary thyroid cancer and tissues. *P < 0.05; **P < 0.01

Fig 5: Competitive binding of KU70 by LSD1 and SIRT1A. Summary of domain mapping of KU70 interaction with LSD1 and SIRT1 in HEK293T cells. Left panel depicts myc-tagged KU70 constructs used along with either wild-type HA-tagged LSD1 (in B) or flag-tagged SIRT1 (in C). Table on the right summarizes relative binding strength for KU70 domains with LSD1 or SIRT1. B, C. Co-IP of myc-KU70 followed by immunoblots of HA-tagged LSD1 (B) or flag-tagged SIRT1 (C) with co-transfected myc-KU70 from HEK293T cells. D. Co-IP of WT or ?SAP myc-tagged KU70 with varying amounts of WT-SIRT1, WT-LSD1 or vector alone in HEK293T cells. Total amount of transfected DNA was held constant by using empty vector DNA to equalize all samples. E. Schematic summary of KU70 domains for regulating interaction with LSD1 and SIRT1. Both SIRT1 and LSD1 bind to the KU70 core domain. KU70 N-terminus strongly represses SIRT1 binding but C-terminal linker region facilitates SIRT1 interaction. KU70 C-terminal SAP motif strongly represses LSD1 binding.