RXRalpha acts as a carrier for TR3 nuclear export in a 9-cis retinoic acid-dependent manner in gastric cancer cells.
Retinoid X receptor (RXR) plays a crucial role in the cross talk between retinoid receptors and other hormone receptors including the orphan receptor TR3, forming different heterodimers that transduce diverse steroid/thyroid hormone signaling. Here we show that RXRalpha exhibits nucleocytoplasmic shuttling in MGC80-3 gastric cancer cells and that RXRalpha shuttling is energy-dependent through a nuclear pore complex (NPC)-mediated pathway for its import and an intact DNA binding domain-mediated pathway for its export. In the presence of its ligand 9-cis retinoic acid, RXRalpha was almost exclusively located in the cytoplasm. More importantly, we also show that RXRalpha acts as a carrier to assist translocation of TR3, which plays an important role in apoptosis. Both RXRalpha and TR3 colocalized in the nucleus; however, upon stimulation by 9-cis retinoic acid they cotranslocated to the cytoplasm and then localized in the mitochondria. TR3 export depends on RXRalpha, as in living cells GFP-TR3 alone did not result in export from the nucleus even in the presence of 9-cis retinoic acid, whereas GFP-TR3 cotransfected with RXRalpha was exported out of the nucleus in response to 9-cis retinoic acid. Moreover, specific reduction of RXRalpha levels caused by anti-sense RXRalpha abolished TR3 nuclear export. In contrast, specific knockdown of TR3 by antisense-TR3 or TR3-siRNA did not affect RXRalpha shuttling. These results indicate that RXRalpha is responsible for TR3 nucleocytoplasmic translocation, which is facilitated by the RXRalpha ligand 9-cis retinoic acid. In addition, mitochondrial TR3, but not RXRalpha, was critical for apoptosis, as TR3 mutants that were distributed in the mitochondria induced apoptosis in the presence or absence of 9-cis retinoic acid. These data reveal a novel aspect of RXRalpha function, in which it acts as a carrier for nucleocytoplasmic translocation of orphan receptors.
Lin
Xiao-Feng
XF
Key Laboratory of the Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Sciences, Xiamen University, Xiamen, Fujian Province, 361005, China.
Zhao
Bi-Xing
BX
Chen
Hang-Zhi
HZ
Ye
Xiao-Feng
XF
Yang
Chao-Yi
CY
Zhou
Hai-Ying
HY
Zhang
Ming-Qing
MQ
Lin
Sheng-Cai
SC
Wu
Qiao
Q
D016475:3T3 Cells; D021581:Active Transport, Cell Nucleus / Q000502:physiology; D000077556:Alitretinoin; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation / Q000502:physiology; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000378:metabolism; D022022:Nuclear Pore / Q000378:metabolism; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D021381:Protein Transport / Q000502:physiology; D034622:RNA Interference; D011987:Receptors, Steroid / Q000235:genetics / Q000378:metabolism*; D011988:Receptors, Thyroid Hormone / Q000235:genetics / Q000378:metabolism*; D047490:Retinoid X Receptor alpha / Q000235:genetics / Q000378:metabolism*; D013274:Stomach Neoplasms / Q000378:metabolism*; D014212:Tretinoin / Q000378:metabolism*
D004268:DNA-Binding Proteins; C539821:NR4A1 protein, human; C539818:Nr4a1 protein, mouse; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1; D016376:Oligonucleotides, Antisense; D011987:Receptors, Steroid; D011988:Receptors, Thyroid Hormone; D047490:Retinoid X Receptor alpha; D049452:Green Fluorescent Proteins; D000077556:Alitretinoin; D014212:Tretinoin