Fig. 1. Three RCC1 transcript variants are expressed in humans. (A) Linear representation of RCC1 protein domains (not to scale), showing sequence alignment of the NTR (residues 1-27) of human RCC1 protein with insert-containing RCC1 isoforms from: human (hs); chimpanzee (pt); rhesus monkey (macm); golden hamster (ma); mouse (mm); and African clawed frog (xl). RCC1 homologues in species other than human are not shown. Insert sequences are shown in bold. Phosphorylation sites (serines 2 and 11) are indicated by circles labelled P. (B) Schematic showing how alternative mRNA splicing around and within exon 6' could generate the known RCC1 transcript variants. (C) Isoform-specific antibodies were generated to peptides corresponding to RCC1 sequences. Anti-R-INS should detect insert-containing isoforms, whereas the anti-R-DIS epitope should be disrupted by them. (D) Precipitation of RCC1 proteins from asynchronous HeLa cell extracts by GST-RanT24N or GST as a control, with analysis by immunoblotting with RCC1 isoform-specific antibodies. (E) Immunoblotting of human cultured cell lysates. (F) Immunoblotting of normal human tissue lysates on an Instablot (IMGENEX) membrane (Cambridge Bioscience), which is pre-loaded with SDS-PAGE-separated normal tissue lysates (20 µg protein/lane). The blot was probed using anti-R-INS (upper panel), then re-probed using anti-RCC1 (lower panel). RCC1 isoforms are indicated, with the question mark signifying a possible RCC1 band of unknown identity. Molecular mass (in kDa) is indicated on the left of the immunoblots.