Thyroid hormone receptors (T3Rs) are hormone-regulated transcription factors that play important roles in vertebrate homeostasis, differentiation, and development. T3Rs are synthesized as multiple isoforms that display tissue-specific expression patterns and distinct transcriptional properties. Most T3R isoforms associate with co-activator proteins and mediate transcriptional activation only in the presence of thyroid hormone. The pituitary-specific T3Rbeta-2 isoform departs from this general rule and is able to interact with p160 co-activators, and to mediate transcriptional activation in both the absence and presence of hormone. We report here that this hormone-independent activation is mediated by contacts between the unique N terminus of T3Rbeta-2 and an internal interaction domain in the SRC-1 (steroid receptor co-activator-1) and GRIP-1 (glucocorticoid receptor interacting protein 1) co-activators. These hormone-independent contacts between T3Rbeta-2 and the p160 co-activators are distinct in sequence and function from the LXXLL motifs that mediate hormone-dependent transcriptional activation and resemble instead a mode of co-activator recruitment previously observed only for the steroid hormone receptors and only in the presence of steroid hormone. Our results suggest that the transcriptional properties of the different T3R isoforms represent a combinatorial mixture of repression, antirepression, and hormone-independent and hormone-dependent activation functions that operate in conjunction to determine the ultimate transcriptional outcome.