Figure 3

The H3BD of MTA1 and MTA2 mediates the in vitro binding of H3 tail peptide by NURD. A. Mammalian expressed MTA1(1–460) but not MTA1(454–715) was incorporated into the core HDAC1/2 and RbAp46/48 containing complex. The MTA1 and its N- and C-terminal regions were expressed in 293T cells and assayed for association with other subunits of NURD complex by IP-western analysis. Note that the N-terminal but not the C-terminal region of MTA1 associated with HDAC1/2 and RbAp48 and that no association with CHD4 was detected for MTA1. Input, 10%. B. The MTA1 C-terminal region but not the N-terminal region that was incorporated into the HDAC1/2-RbAp46/48 core complex bound the H3 tail peptide in in vitro pulldown assay. The samples in A were subjected to in vitro pulldown assay. Note that no binding was detected for Flag-MTA1(1–460), whereas Flag-MTA1(454–715) bound the H3 tail peptide as efficient as Flag-MTA1. Input, 10%. C. The C-terminal region of MTA1 binds H3 tail peptide independent of HDAC1/2 and RbAp48. The 293T-expressed, Flag-tagged MTA1, MTA1(1–460) and MTA1(454–715) were purified as illustrated in the top panel and then subjected to in vitro pulldown assay for binding of H3 tail peptide. D. Mammalian expressed MTA2(1–434) but not MTA2(427–668) was incorporated into the endogenous NURD complex. The MTA2 and its N- and C-terminal regions were expressed in 293T cells and assayed for association with other subunits of NURD complex by IP-western analysis. Note that the N-terminal but not the C-terminal region of MTA2 associated with HDAC1/2 and CHD4. Input, 10%. E. The C-terminal region but not the N-terminal region that was incorporated into the endogenous NURD complex bound the H3 tail peptide in in vitro pulldown assay. The samples in D were subjected to in vitro pulldown assay. Input, 10%.