Figure 3

Subcellular localization of HML-2 SP fusion protein. (A) Schematic representation of SP fusion proteins. N-extensions of HML-2 SP of different length (aa 1–75; aa 1–94; aa 1–96) were cloned in frame with the monomeric red fluorescent protein (mRFP). NLS: putative nuclear localization signal; NES: putative nuclear export signal; h: hydrophobic core. (B) Western blot analysis of HeLa cells transiently expressing mRFP and SP₇₅-RFP. The Western blot was stained with anti-mRFP antibody. mRFP-expressing cells produce mRFP with an approximate molecular weight of 30 kDa. An 18–19 kDa proteolytic product can also be is detected. An SP₇₅-RFP construct produces SP-RFP fusion protein and (RFP) degradation products. (C) Confocal sections showing SP₇₅-RFP, SP₉₆-RFP and SP₉₄-RFP fluorescence in red and co-immunostained nucleolar markers B23/nucleophosmin or C23/nucleolin in green. The lower right panel shows GFPcORF/Rec fluorescence in green and nucleolar markers in red. Co-localization of proteins is indicated in yellow. Images show HeLa cells fixed 24 hours post-transfection. White bar = 10 μm.