T3 is required for normal early development, but relatively few T3-responsive 
target genes have been identified. In general, in vitro stem cell 
differentiation techniques stimulate a wide range of developmental programs, 
including thyroid hormone receptor (TR) pathways. We developed several in vitro 
stem cell models to more specifically identify TR-mediated gene expression in 
early development. We found that embryonic carcinoma (EC) cells have reduced T3 
nuclear binding capacity and only modestly express the known T3 target genes, 
neurogranin (RC3) and Ca2+/calmodulin-dependent protein kinase IV (CaMKIV), in 
response to T3. Full T3 induction in transient transfection of EC cells was 
restored with cotransfection of a TR expression vector. We, therefore, performed 
gene expression profiles in wild-type embryonic stem (ES) cells compared with 
expression in cells with deficient (EC) or mutant TR (TRalpha P398H mutant ES 
cells), to identify T3 target genes. T3 stimulation of wild-type ES cells 
altered mRNA expression of 610 known genes (26% of those studied), although only 
approximately 60 genes (1%) met criteria for direct T3 stimulation based on the 
magnitude of induction and requirement for the presence of TR. We selected five 
candidate T3 target genes, neurexophilin 2, spermatid perinuclear RNA-binding 
protein (SPNR), kallikrein-binding protein (KBP), prostate-specific membrane 
antigen (PSMA), and synaptotagmin II, for more detailed study. T3 responsiveness 
of these genes was evaluated in both in vitro endogenous gene expression and in 
vivo mouse model systems. These genes identified in a novel stem cell system, 
including those induced and repressed in response to T3, may mediate thyroid 
hormone actions in early development.