The trans‐inhibitory Rep78 protein of adeno‐associated virus binds to TAR region DNA of the human immunodeficiency virus type 1 long terminal repeat

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Abstract
]he large rep gene products, Rep78 and Rep68, of adeno‐associated virus (AAV) are pleiotropic effector proteins which are required for AAV DNA replication and the trans‐regulation of AAV gene expression. Apart from these essential functions prerequisite for the life cycle of AAV, these rep products are able to inhibit the replication and gene expression of human immunodeficiency virus type 1 (HIV‐1) and a number of DNA viruses. Here, it is demonstrated that Rep78, as a chimeric with the maltose binding protein, directly binds the full‐length HIV‐1 long terminal repeat (LTR), and to a subset of these sequences containing the trans‐activation response (TAR) sequence as DNA. These interactions, an effector protein physically binding a target promoter, suggest a direct mechanism of action for Rep78 inhibition. Furthermore, competitive binding studies between the TAR region and the full‐length HIV‐LTR, strongly suggested that another site(s) within the LTR was also bound by Rep78. Finally, as Rep78 binding is also believed to be affected by secondary structure within the DNA, it was found that Rep78 preferentially binds with HIV‐LTR sequences with promoted secondary structure generated by heat denaturation and rapid cooling.