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Figure 2 | BMC Biotechnology

Figure 2

From: Novel system uses probasin-based promoter, transcriptional silencers and amplification loop to induce high-level prostate expression

Figure 2

Mechanism of action of the Ad/GFP DiSTRES vector. Mechanism of action of the Ad/GFPDiSTRES vector when transduced into a prostate cell (top panel). We hypothesize that the following sequence of events would occur in the prostate cell: (1) Both the tTA and the LacI genes would be induced, as a result of activation of the prostate specific TRE-ARR2PB promoters that drive the expressions of these genes; (2) Expression of tTA would then lead to transcriptional activation of the gene of interest (e.g., GFP), through binding of tTA to the TRE promoter; (3) The tTA protein would also bind to the TRE within its own TRE-ARR2PB promoter, thereby activating further transcription of itself and therefore establishing a positive feedback amplification loop of tTA expression; (4) Meanwhile, the LacR protein would bind to the LacR-responsive promoter of the tTS gene, thereby leading to suppression of tTS gene transcription; and (5) Finally, the tTA-driven amplification loop would lead to further expression of both the gene of interest and the LacR (and subsequently further repress the tTS). Mechanism of action in non-prostate cells (bottom panel). In the non-prostate cell, we propose the following scenario: (1) Because the TRE-ARR2PB promoter is prostate-specific, neither the tTA nor the LacR proteins would be expressed; (2) In the absence of the LacR protein, the LacR-responsive promoter acts essentially like its constitutively active parental promoter (the CMV promoter). As a result, the tTS gene is now actively transcribed; and (3) Expression of the tTS protein would then lead to the transcriptional silencing of all TRE-regulated promoters, and therefore, the gene of interest (GFP), tTA and LacR would all be suppressed upon binding of the tTS to the TRE's within their respective promoters. Note: Tetracycline and/or doxycycline are not added to this system. Adding these drugs would act to negate the effects of the differential regulation of tTA versus tTS. The result would be basal expression of GFP in both cell types, and therefore, a loss in prostate-specificity.

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