I factors in Drosophila melanogaster are non-LTR retrotransposons that transpose at very high frequencies in the germ line of females resulting from crosses between reactive females (devoid of active I factors) and inducer males (containing active I factors). Constructs containing I factor ORF1 under the control of the hsp70 promoter repress I factor activity. This repressor effect is maternally transmitted and increases with the transgene copy number. It is irrespective of either frame integrity or transcriptional orientation of ORF1, suggesting the involvement of a homology-dependent trans-silencing mechanism. A promoterless transgene displays no repression. The effect of constructs in which ORF1 is controlled by the hsp70 promoter does not depend upon heat-shock treatments. No effect of ORF1 is detected when it is controlled by the I factor promoter. We discuss the relevance of the described regulation to the repression of I factors in I strains.