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Fig. 8. TBX5 and TBX20 physically interact. Cell lysates containing GST- and/or HA-tagged proteins were incubated on GST and eluted, and separated by SDS-PAGE. GST proteins were detected using anti-GST antibodies and HA-tagged proteins were detected with anti-HA antibodies. (A) Association of TBX5 with TBX20 is shown by pull-down of HA-TBX20 with GST-TBX5. HA-NKX2-5 serves as positive control. Fifteen percent of output and 7.5% of input was probed. (B,C) Pull-down of full-length HA-TBX20 with a GST-tagged TBX5 deletion series reveals an interaction domain in the N terminus and T-box region of TBX5. Each reaction was probed with anti-HA antibodies (B). Fifteen percent of output and 7.5% of input was probed. (D,E) Pull-down of HA-TBX20 deletion series with full-length GST-TBX5 reveals an interaction domain within the N-terminus and T-box of TBX20. Each reaction was probed with anti-HA antibodies (D). Fifteen percent of output and 7.5% of input was probed, except in the case of {Delta}N/C, in which the amount of protein probed was only 4% owing to the increase in total amount of {Delta}N/C protein used in pull-down (see text).