ER-based double iCre fusion protein allows partial recombination in forebrain.
Here we describe the generation of a new tamoxifen-inducible double Cre fusion protein generated by fusing two ERT2 domains onto both ends of the iCre recombinase (a codon improved Cre recombinase). This Cre fusion protein (ERiCreER) had a twofold increased activity in cell culture assays than the previously described MerCreMer Cre double fusion protein. ERiCreER was targeted to the brain by placing it under the control of the promoter from the CamKIIalpha gene using a 170 kb BAC. The fusion protein was detected in hippocampus, cortex, striatum, thalamus, and hypothalamus but not in cerebellum. The ERiCreER was cytoplasmatic in the absence of tamoxifen and translocated into the nucleus upon tamoxifen administration. The activity of the ERiCreER was tested in vivo by mating the CamKIIalpha ERiCreER transgenic line with mice harbouring exon 10 of the CREB gene flanked by two LoxP sites. In the absence of tamoxifen, no background activity was detected in mice older than 6 months. After tamoxifen administration, most if not all of the ERiCreER fusion protein translocated from the cytoplasm to the nucleus; however, only 5-10% of the "floxed" CREB allele was recombined. Recombination was also visualised at the cellular level by following the upregulation of the CREM protein, which corresponds precisely with CREB loss/recombination. Unlike in other tissues (Sohal et al., 2001; Tannour-Louet et al., 2002), it appears that in brain, although ERiCreER can bind tamoxifen, the Cre-recombinase cannot be fully activated.
Casanova
Emilio
E
Deutsches Krebsforschungszentrum, Heidelberg, Germany. e.casanova@dkfz.de
Fehsenfeld
Sandra
S
Lemberger
Thomas
T
Shimshek
Derya R
DR
Sprengel
Rolf
R
Mantamadiotis
Theo
T
D000818:Animals; D001921:Brain / Q000378:metabolism / Q000502:physiology*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D019426:Integrases / Q000235:genetics*; D051379:Mice; D008822:Mice, Transgenic; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D011995:Recombination, Genetic / Q000502:physiology*; D013629:Tamoxifen / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics*
D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D013629:Tamoxifen; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C045073:Cre recombinase; D019426:Integrases