Subtype selective tetracycline agonists and their application for a two-stage regulatory system.
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Doxycycline↗
Biomedical subjects
Lars Drueppel
Publications and source records attributed to Lars Drueppel.
An optimized conditional suicide switch using doxycycline-dependent expression of human tBid.
We have examined in a HeLa model system whether tight doxycycline-dependent expression of human tBid can be used as a fast and efficient suicide switch. A stably transfected cell line expressing human tBid in a strictly dox-dependent manner showed fast and efficient killing with up to 98 % specific cell death 24 h after induction. The survival rate was only 0.6 % after eleven days of dox treatment. Very low amounts of tBid were sufficient to induce apoptosis indicating that stringent control of the dox-dependent suicide gene is essential for cell survival in the absence of inducer. Using human tBid, an endogenous effector protein with low immunogenic potential, and doxycycline, an inducer with low cytotoxicity and a long record of safe use in humans, this conditional suicide switch provides a tool for various applications, e.g., adoptive immune transfer.
Apoptosis↗
Inducible DNA-loop formation blocks transcriptional activation by an SV40 enhancer.
It is well established that gene expression in eukaryotes is controlled by sequence-dependent binding of trans-acting proteins to regulatory elements like promoters, enhancers or silencers. A less well understood level of gene regulation is governed by the various structural and functional states of chromatin, which have been ascribed to changes in covalent modification of core histone proteins. And, much on how topological domains in the genome take part in establishing and maintaining distinct gene expression patterns is still unknown. Here we present a set of regulatory proteins that allow to reversibly alter the DNA structure in vivo and in vitro by adding low molecular weight effectors that control their oligomerization and DNA binding. Using this approach, we completely regulate the activity of an SV40 enhancer in HeLa cells by reversible loop formation to topologically separate it from the promoter. This result establishes a new mechanism for DNA-structure-dependent gene regulation in vivo and provides evidence supporting the structural model of insulator function.
Base Sequence↗
A short autonomous repression motif is located within the N-terminal domain of CTCF.
The vertebrate transcription factor CTCF is not only involved in transcriptional activation, insulation and genomic imprinting, but also in transcriptional repression. Sequence motifs mediating these activities have not been identified so far. We have mapped a short repression motif to residues 150-170 within the N-terminal domain of CTCF. This motif is active in HeLa, HEK293 and COS-7 cell lines where it is both sufficient and necessary for silencing either an SV40-, or a CMV-enhancer. It also represses the basal activity of an SV40 core promoter. Since this autonomous repression motif displays no sequence similarity to any other regulatory protein, it represents a yet unknown co-repressor recruiting motif.
Animals↗