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. 1999 Jan 19;96(2):709–714. doi: 10.1073/pnas.96.2.709

Figure 1.

Figure 1

Interaction between RHA and RRE. (a) In vitro binding of RHA to CTE but not to RRE in the presence or absence of Rev. RNA Selection with biotin-labeled RRE (lanes 2, 3, and 6) or CTE (lane 4) was carried out as described (13). Recombinant Rev was added to the nuclear extract in some samples (lanes 2, 5, and 6). Lanes 1–4 were probed with anti-RHA antibodies and lanes 5–6 were probed with anti-Rev antibodies. Lane 1 (nuclear extract) and lane 5 (Rev mixed with nuclear extract) were positive controls for direct immunodetection with antibodies without RNA selection. The arrow indicates the position of the Rev-specific band in lanes 5 and 6. (b) Detection of in vivo interactions between RHA and RRE or Rev-RRE by using immunoprecipitation and RT-PCR. Lysates from cells transfected with pRev and pRRE (lanes 1–6) or pRRE alone (lanes 7–12) were immunoprecipitated with normal serum (lanes 1, 2, 7, and 8), anti-Rev antibodies (lane, 3, 4, 9, and 10), or anti-RHA antibodies (lanes 5, 6, 11, and 12). Half of the RNA obtained from the precipitate was subjected to RT (lanes 1, 3, 5, 7, 9, and 11) and PCR amplification using RRE-specific primers. The other half was processed under the same conditions without RT (lanes 2, 4, 6, 8, 10, and 12). The expected product for RRE was 230 bp in length.