Abstract: RNA molecules have been shown to play a variety of functional roles in biological processes. RNA sequences can adopt 3-D conformations able to regulate transcription. RNA-based transcriptional regulators when recruited to a DNA template act to promote or suppress the transcription of nearby genes. The regulators are thought to act by mimicing the activation domains of protein transcriptional activators. These RNA-based transcriptional regulators may be engineered to regulate transcription based on the binding of a ligand such as a small molecule. RNA sequences of the regulators may be evolved to produce regulators with a greater degree of transcriptional activation or suppression. The RNA-based system of the present invention is useful in studying biological processes and in altering biological process in a therapeutic context.

Abstract: Ligand-dependent inteins allow for modulation of a protein's activity in vivo. Upon binding of the ligand to the ligand-dependent intein inserted into the protein of interest, the hybrid protein undergoes protein splicing removing the intein. The activity of the spliced protein is then restored. A 4-hydroxytamoxifen-dependent intein based on the M. tuberculosis RecA intein is prepared and demonstrated in a variety of exteins contexts. The invention provides a system for engineering other ligand-dependent inteins and using them, including the ligand-dependent inteins themselves, hybrid proteins with the inserted ligand-dependent inteins, polynucleotides encoding inteins and hybrid proteins, and engineered cells. Kits with the materials and reagents necessary for preparing and using ligand-dependent inteins are also included.

Abstract: Ligand-dependent inteins allow for modulation of a protein's activity in vivo. Upon binding of the ligand to the ligand-dependent intein inserted into the protein of interest, the hybrid protein undergoes protein splicing removing the intein. The activity of the spliced protein is then restored. A 4-hydroxytamoxifen-dependent intein based on the M. tuberculosis RecA intein is prepared and demonstrated in a variety of exteins contexts. The invention provides a system for engineering other ligand-dependent inteins and using them, including the ligand-dependent inteins themselves, hybrid proteins with the inserted ligand-dependent inteins, polynucleotides encoding inteins and hybrid proteins, and engineered cells. Kits with the materials and reagents necessary for preparing and using ligand-dependent inteins are also included.