Single-chain Tet transregulators.
We demonstrate here that the Tet repressor (TetR), a dimeric allosterical regulatory protein, can be converted to a fully functional monomer when connected by a 29 amino acid linker. TetR-based transregulators are widely used to regulate gene expression in eukaryotes. They can be fused to form single-chain (sc) Tet transregulators with two TetR moieties and one eukaryotic regulatory domain. Sc variants of transactivator and transsilencer exhibit the same regulatory properties as their respective dimeric counterparts in human cell lines. In particular, the reverse 'tet-on' phenotype of rtTA variants is also present in the sc variants. Coexpression of a reverse transactivator and sc transsilencer leads to reduced background expression and shows full activation upon induction. The data demonstrate that sc Tet transregulators exhibit the phenotype of their respective dimers and lack functional interference when coexpressed in the same cell.
Krueger
Christel
C
Lehrstuhl für Mikrobiologie, Friedrich-Alexander Universität Erlangen-Nürnberg, Staudtstrasse 5, D-91058 Erlangen, Germany.
Berens
Christian
C
Schmidt
Andreas
A
Schnappinger
Dirk
D
Hillen
Wolfgang
W
D002460:Cell Line; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism; D020868:Gene Silencing; D006367:HeLa Cells; D006801:Humans; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D015534:Trans-Activators / Q000037:antagonists & inhibitors
D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D015534:Trans-Activators; C034811:tetracycline resistance-encoding transposon repressor protein