Transcription artificial

Disubstituted4-dimethylaminopyridines as artificial inhibitors of transcription factors involved in HIV replication 97YGK697. 

PIAS Proteins. Figure 1 Reporter gene assay with an interferon-y-driven artificial promoter demonstrating the inhibitory transcriptional activity of increasing amounts of PIAS on STAT1 signal transduction. 

Reverse transcription is the copying of an RNA molecule back into its DNA complement. The enzymes that perform this function are called reverse transcriptases. Reverse transcription is used naturally by retroviruses to insert themselves into an organism s genome. Artificially induced reverse transcription is a useful technique for translating unstable messenger RNA (mRNA) molecules into stable cDNA. 

Fig. 3.17 Activation of gene transcription by artificial transcription factors. (Top) The artificial activator is composed of three separate functional domains. The DNA binding domain consists of the pyrrole/imidazole polyamides (shown as connected arrows). A tethered linker domain (shown as a coil) connects the DNA binding domain to the peptide activation domain (AD, shown as an oval).

Upon binding, the artificial transcription factor recruits the necessary transcriptional machinery for gene activation. (Bottom, left) Ball-and-stick model for a polyamide conjugated to the VP2 activation domain. Symbols are as in Fig. 3.4. (Bottom, right) Structure of the polyamide-VP2 conjugate with the polyproline linker domain in brackets... 

P.B. Dervan, and M. Ptashne. Towards a minimal motif for artificial transcriptional activators. Chem. Biol. 2001, 8, 583—592. 

Arora, P.S., A.Z. Ansari, T. P. Best, M. Ptashne, and P. B. Dervan. Design of artificial transcriptional activators with rigid poly-L-proline linkers. J. Am. Chem. 

M0LLEGAARD N.E., BuCHARDT O., Egholm M., Nielsen P.E. Peptide nucleic acid-DNA strand displacement loops as artificial transcription promoters. Proc. Natl Acad. Sci. USA 1994 91 3892-3895. 

The participation of the nuclear receptors in the machinery of gene transcription takes place by means of specific domains of the molecule known as transactivators (abbreviation for transcription activators). These are made up of sequences of amino acids that interact by means of protein-protein contacts with other transcription factors. The artificial alteration of these sequences has as a consequence the inability of the hormone to induce gene expression (Beato et al. 1996 Klug et al. 1987 Lones et al. 1995). 

The lac operon is ordinarily subject to repression and is activated by the presence of an inducer, now known to be allolactose, D-Galp-(il— 6-d-G1c. However, in experimental work artificial inducers such as isopropyl-[5-o-thiogaIactoside (IPTG) are most often used. Jacob and Monod postulated that the free repressor protein binds to the operator. In the presence of the inducer a conformational change takes place, destroying the affinity of the repressor protein for the operator site. Thus, in the presence of inducer the operator is not blocked, and the transcription takes place. Such an operon is said to be negatively controlled and inducible. 

The DNA binding ability of peptide nanostructures has also been reported by the artificial dimerization of peptide sequences corresponding to the contact region of the transcriptional activator protein GCN4. Cuenoud and Schepartz... 

Fig. 7.1. A planarian individual of a line transformed with a Hermes transposon-derived vector 12 months after microinjection and subsequent electroporation. Besides the transposon part, the vector contained the E(enhanced)GFP reporter gene under the control of an artificial promoter that responds to the transcription factor Pax6 (Gonzalez-Estevez et ai, 2003). Binding sites for this transcription factor are found in rhodopsin and other photoreceptor-specific genes. Reporter-gene activity was found in the eyes (arrows) of transformed individuals or in regenerated heads as a mosaic of EGFP-positive photoreceptor cells. Figure courtesy of Dr E. Salo.

Artificially enhanced error rates needed for the creation of sequence diversity in a population can be achieved readily with PCR. Reverse transcription and transcription are also susceptible to increase of mutation rates. These two and other new techniques for RNA amplification provide universal and efficient tools for the study of molecular evolution under laboratory conditions and make the usage of viral replicases with their undesirable sequence specificities obsolete. 

Dreier, B., Beerli, R.R., Segal, D.J., Flippin, J.D. Barbas C.F., 3rd. Development of zinc finger domains for recognition of the 5 -ANN-3 family of DNA sequences and their use in the construction of artificial transcription factors. J Biol Chem 2001 276 29466-29478. 

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