Mediator subunit MED1 modulates intranuclear dynamics of the thyroid hormone receptor

• Published in: Journal of cellular biochemistry Vol. 121; no. 4; pp. 2909 - 2926
• Main Authors: Femia, Matthew R., Evans, Rochelle M., Zhang, Jibo, Sun, Xiaopeng, Lebegue, Caroline J., Roggero, Vincent R., Allison, Lizabeth A.
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01.04.2020
• Subjects: Binding sites; Distribution patterns; Domains; Epidermal growth factor; Exports; Fluorescence; Fluorescence recovery after photobleaching; fluorescence recovery after photobleaching (FRAP); Gene expression; Growth factors; intranuclear mobility; Localization; Mediator subunit 1 (MED1); Mediator subunit 13 (MED13); Mobility; Nuclear transport; Phosphorylation; Photobleaching; Receptors; Retention; Thyroid; Thyroid gland; thyroid hormone receptor; Thyroid hormone receptors; Triiodothyronine; Mediator subunit 13 (MED13); fluorescence recovery after photobleaching (FRAP); thyroid hormone receptor; intranuclear mobility; Mediator subunit 1 (MED1)
• ISSN: 0730-2312; 1097-4644
• DOI: 10.1002/jcb.29532

The thyroid hormone receptors (TRs) mediate thyroid hormone (T3)‐dependent gene expression. The nuclear import and export signals that direct TR shuttling are well characterized, but little is known about factors modulating nuclear retention. We used fluorescence‐based nucleocytoplasmic scoring and fluorescence recovery after photobleaching in transfected cells to investigate whether Mediator subunits MED1 and MED13 play a role in nuclear retention of TR. When MED1 was overexpressed, there was a striking shift towards a greater nuclear localization of TRβ1 and the oncoprotein v‐ErbA, subtypes with cytosolic populations at steady‐state, and TRβ1 intranuclear mobility was reduced. For TRα1, there was no observable change in its predominantly nuclear distribution pattern or mobility. Consistent with a role for MED1 in nuclear retention, the cytosolic TRα1 and TRβ1 population were significantly greater in MED1−/− cells, compared with MED1+/+ cells. Exposure to T3 and epidermal growth factor, which induces MED1 phosphorylation, also altered TR intranuclear dynamics. Overexpression of miR‐208a, which downregulates MED13, led to a more cytosolic distribution of nuclear‐localized TRα1; however, overexpression of MED13 had no effect on TRβ1 localization. The known binding site of MED1 overlaps with a transactivation domain and nuclear export signal in helix 12 of TR's ligand‐binding domain (LBD). Coimmunoprecipitation assays demonstrated that TR's LBD interacts directly with exportins 5 and 7, suggesting that binding of exportins and MED1 to TR may be mutually exclusive. Collectively, our data provide evidence that MED1 promotes nuclear retention of TR, and highlight the dual functionality of helix 12 in TR transactivation and nuclear export. By altering Mediator subunit levels in cells transfected with fluorescent fusion proteins, we show that MED1 promotes nuclear retention and reduces intranuclear mobility of the thyroid hormone receptor.