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. 2003 Apr;23(8):2981–2990. doi: 10.1128/MCB.23.8.2981-2990.2003

FIG. 3.

FIG. 3.

Localization of ASAP complex components in HeLa cells. (a) HeLa cells were transiently transfected with a Flag-RNPS1 expression construct. Overexpression of Flag-RNPS1 leads to formation of a few large mega-speckles (left panel, arrowheads) or more abundant and smaller speckles (right panel). Localization of Flag-RNPS1 and SAP18 was analyzed by indirect immunofluorescence with antibodies against the Flag tag and SAP18. SAP18 is present in multiple nuclear sites in untransfected cells (left panel, arrows) but colocalizes with Flag-RNPS1 in transfected cells. (b) HeLa cells were transfected and analyzed with antibodies against the Flag tag and Acinus as in panel a. Mega-speckles caused by overexpression of Flag-RNPS1 are indicated by arrowheads, and localization of Acinus in nuclear speckles in untransfected cells is indicated by arrows. Acinus colocalizes with Flag-RNPS1 in transfected cells. (c) HeLa cells transiently transfected with Flag-RNPS1 were analyzed with a fluorescein isothiocyanate-coupled monoclonal antibody against the Flag tag and by indirect immunofluorescence with an antibody against Sin3. Sin3 shows nuclear staining and does not colocalize with Flag-RNPS1 in the mega-speckles. (d) Untransfected HeLa cells were analyzed for localization of SAP18 and Acinus by indirect immunofluorescence with antibodies against SAP18 and the carboxyl terminus of Acinus, respectively. SAP18 and Acinus colocalize in nuclear speckles. (e) Untransfected HeLa cells were analyzed as in panel d with antibodies against SAP18 and SC35, respectively. SAP18 localizes to nuclear speckles containing SC35.