![Fig. 2. Identification of MLL processing sites. / (A) Purification of MLL proteins. BOSC23 cells were transfected with an MLL-expression vector [pLNCX MLL(H)], lysed 48 hours later, and subjected to immunoprecipitation with an anti-HA antibody 3F10. An aliquot of the immunoprecipitate was separated in a denaturing gel and stained by the silver stain method. Full-length MLL, the purified C-terminal 180-kDa fragment (MLLC), and the copurified 300-kDa band are indicated. Migration of molecular weight markers is indicated on the left. (B) Amino acid sequences of MLL processing sites. The sequences around the 2 processing sites of human MLL (hMLL) and putative processing sites of mouse MLL (mMLL), pufferfish MLL (fMLL), Drosophila TRX (dTRX), and human MLL2 (hML2) are aligned.12254041 Conserved amino acids are shaded and marked with an asterisk. The processing site is indicated by an arrow in the consensus sequence below. The numbers refer to positions relative to the first amino acid of each protein. (C) Inhibition of processing by mutations in the cleavage sites. BOSC23 cells were transfected with expression constructs encoding proteins indicated above the gel lanes and harvested 48 hours after transfection. All MLL proteins were tagged with HA at their C-termini. The cell lysates were analyzed by immunoblot with anti-HA antibody (left panel) or anti-MLL N-terminal antibody (right panel).](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/100/10/10.1182_blood-2002-04-1015/4/h82223465002.jpeg?Expires=1726822217&Signature=Yl4QOjauyUL6021-aySfgXLGI16vxwP2GSHS3XWY9nyA~TdtTC7lkd98mSu8PvNICXh~NlSdpwVcLRj~D5cAnFTkwperT~HUJMdiDEYzR9ohxnBP2fmiEAA-y7X4C5yorCqFQfNdwTt8tsDFpHjPLmGk3xBVlCfq8oqKGqZ6upgG0fln-sQ-UnvV3LJHGOGxk-zMghkqufMRMQqzLE636jpWkZdq7-LW6tdy6mJApKEcDC1NHfswcQ0tSDM~NTxXoav2t3I1JEdcBxW~~~glC0nSUTyrlZ1s3Qrm8oSvwVQHPs1rLFDWGHZ~9MoJlnA6adx~gxF2V~ACMS9mmPEyew__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Identification of MLL processing sites.
(A) Purification of MLL proteins. BOSC23 cells were transfected with an MLL-expression vector [pLNCX MLL(H)], lysed 48 hours later, and subjected to immunoprecipitation with an anti-HA antibody 3F10. An aliquot of the immunoprecipitate was separated in a denaturing gel and stained by the silver stain method. Full-length MLL, the purified C-terminal 180-kDa fragment (MLLC), and the copurified 300-kDa band are indicated. Migration of molecular weight markers is indicated on the left. (B) Amino acid sequences of MLL processing sites. The sequences around the 2 processing sites of human MLL (hMLL) and putative processing sites of mouse MLL (mMLL), pufferfish MLL (fMLL), Drosophila TRX (dTRX), and human MLL2 (hML2) are aligned.1,2,25,40 41 Conserved amino acids are shaded and marked with an asterisk. The processing site is indicated by an arrow in the consensus sequence below. The numbers refer to positions relative to the first amino acid of each protein. (C) Inhibition of processing by mutations in the cleavage sites. BOSC23 cells were transfected with expression constructs encoding proteins indicated above the gel lanes and harvested 48 hours after transfection. All MLL proteins were tagged with HA at their C-termini. The cell lysates were analyzed by immunoblot with anti-HA antibody (left panel) or anti-MLL N-terminal antibody (right panel).
