Could you please expand on the isoform issue? That is, which isoforms do(does) the Genentech antibody not react with? Thanks!
Maybe these investigators have some insights?? Perhaps a phone call would yield...... "it was a no brainer, it wasn't going to work" and some rationale???.......
Biochem Pharmacol 2002 Sep;64(5-6):919
Designed transcription factors as tools for therapeutics and functional genomics.
Urnov F, Rebar E.
Sangamo Biosciences, Inc., Pt. Richmond Tech. Center, 501 Canal Blvd., Suite A100, 94804, Richmond, CA, USA
The paucity of tools that control expression of specific genes in vivo represents a major limitation of functional genomics in mammals; most available small-molecule regulators of transcription-e.g. histone deacetylase inhibitors-exert pan-genomic effects. Recent developments in understanding the role of chromatin in regulating the genome, and of protein-DNA interactions have allowed the development of designed transcription factors that regulate specific genes in vivo (Reik et al., Curr Opin Genet Dev 2002;12:233). These proteins contain two modules: (i) a zinc finger protein (ZFP)-based DNA-binding domain (DBD) designed to recognize a specific sequence (for example, a motif in the promoter of a certain gene); (ii) a functional module (for example, a transcriptional activation or repression domain). Recent data describe the use of such designed transcription factors to regulate a variety of clinically relevant gene targets in human cells: these include MDR1, erythropoietin, erbB-2 and erbB-3, VEGF, and PPARgamma. In the case of VEGF (Liu et al., J Biol Chem 2001;276:11323), proportional upregulation by the designed transcription factor of all three distinct splice isoforms generated by this locus was observed, illuminating the utility of endogenous gene control in therapeutic settings (proper isoform ratio is essential for the proangiogenic function of VEGF). In the case of PPARgamma, use of a transcriptional repressor designed to downregulate the expression of two PPARgamma isoforms allowed "mutation-free reverse genetics" analysis that illuminated a unique role for the PPARgamma2 isoform in adipogenesis (Ren et al., Genes Dev 2002;16:27). The ability to selectively activate or repress specific mammalian genes in vivo using designed transcription factors thus has considerable promise in clinical and in basic science settings. |
