Mice heterozygous for the Nxph4-2A-CreERT2-D targeted mutation are viable and fertile. The Nxph4-2A-CreERT2-D targeted mutation has a viral 2A oligopeptide (T2A) that mediates ribosomal skipping and a CreERT2 fusion gene (CreERT2 fusion protein) inserted downstream of the neurexophilin 4 translational STOP codon.
As such, Nxph4-2A-CreERT2-D mice have both endogenous gene and CreERT2 fusion protein expression directed to Nxph4-expressing cells by the endogenous promoter/enhancer elements of the neurexophilin 4 locus. CreERT2 fusion gene activity is inducible; observed following tamoxifen administration. When Nxph4-2A-CreERT2-D mice are bred with mice containing loxP-flanked sequences, tamoxifen-inducible Cre-mediated recombination will result in deletion of floxed sequences in the Nxph4-expressing cells of the double mutant offspring.
Specifically, the donating investigator reports that following tamoxifen induction, Nxph4-2A-CreERT2-D directs reporter gene expression to cortical layer 6b neurons and in other scattered small cells (possibly glia) throughout the cortex, in a pattern that partially recapitulates the endogenous Nxph4 expression.
The donating investigator reports no CreERT2 activity (no reporter gene expression) is observed prior to tamoxifen exposure.
The donating investigator did not examine CreERT2 activity in tissues other than brain, and did not attempt to generate homozygous mice to date (July 2013).
For characterization information, see images at the Allen Institute for Brain Science website (Nxph4-2A-CreERT2 images).
The Cre-ERT2 fusion protein consists of Cre recombinase fused to a triple mutant form of the human estrogen receptor which does not bind its natural ligand (17β-estradiol) at physiological concentrations but will bind the synthetic estrogen receptor ligands 4-hydroxytamoxifen (OHT or tamoxifen) and, with lesser sensitivity, ICI 182780. Restricted to the cytoplasm, Cre-ERT2 can only gain access to the nuclear compartment after exposure to tamoxifen. To counteract the mixed estrogen agonist effects of tamoxifen injections, which can result in late fetal abortions in pregnant mice, progesterone may be coadministered.
