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Mol Cell Biol. 1986 August; 6(8): 2884-2894

Negative and positive regulation in trans of gene expression from adeno-associated virus vectors in mammalian cells by a viral rep gene product.

J D Tratschin, J Tal and B J Carter

ABSTRACT

We previously described use of the human parvovirus, adeno-associated virus (AAV), as a vector for transient expression in mammalian cells of the gene for chloramphenicol acetyltransferase (CAT). In the AAV vector, pTS1, the CAT gene is expressed under the control of the major AAV promoter p40. This promoter is embedded within the carboxyl-terminal region of an open reading frame (orf-1) which codes for a protein (rep) required for AAV DNA replication. We show here that the rep product has additional trans-acting properties to regulate gene expression. First, deletion or frame-shift mutations in orf-1, which occurred far upstream of p40, increased expression of CAT in human 293 (adenovirus-transformed) cells. This increased CAT expression was abolished when such mutant AAV vectors were transfected into 293 cells together with a second AAV vector which could supply the wild-type AAV rep product in trans. Thus, an AAV rep gene product was a negative regulator, in trans, of expression of CAT in uninfected 293 cells. In adenovirus-infected 293 cells, the function of the AAV rep product was more complex, but in some cases, it appeared to be a trans activator of the expression from p40. In HeLa cells, only trans activation by rep was seen in the absence or presence of adenovirus. Neither activation nor repression by the rep product required replication per se of the AAV vector DNA. Thus, trans-acting negative or positive regulation of gene expression by the AAV rep gene is modulated by factors in the host cell and by the helper adenovirus.

Mol Cell Biol. 1986 August; 6(8): 2884-2894




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