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. 2004 Jan 29;32(2):611–622. doi: 10.1093/nar/gkh223

Figure 4.

Figure 4

YB-1 binds to the 5′-UTR region of YB-1mRNA. (A) Endogenous YB-1 protein binds to the 5′-UTR region of YB-1 mRNA in the cytoplasm. The indicated amounts of KB3-1 and H1299 cell lysate were incubated with 32P-labeled YB-1 5′-UTR RNA at 25°C for 15 min. An aliquot of 2 µg YB-1-specific antibody (Ab) was added to lanes 5, 6, 11 and 12. An arrow indicates the YB-1/YB-1 5′-UTR RNA complex and a double-headed arrow indicates supershifted complexes. (B) Purified YB-1 binds to YB-1 5′-UTR RNA. The indicated amount of GST or GST–YB-1 fusion was incubated with 32P-labeled YB-1 5′-UTR RNA at 25°C for 15 min. An aliquot of 2 µg YB-1-specific antibody (Ab) was added to lanes 8–11. An arrow indicates the YB-1/YB-1 5′-UTR RNA complex and a double-headed arrow indicates supershifted complexes. (C) Rabbit YB-1 (p50) binds to the 5′-UTR region of YB-1 mRNA in the in vitro translation system. The indicated amounts of rabbit reticulocyte lysate (RRL) were incubated with 32P-labeled YB-1 5′-UTR RNA at 25°C for 15 min. An aliquot of 2 µg YB-1-specific antibody (Ab) was added to lanes 4 and 5. An arrow indicates the rabbit YB-1/YB-1 5′-UTR RNA complex and a double-headed arrow indicates supershifted complexes. YB-1/YB-1 5′-UTR RNA probe complexes were separated as described above.