The SUMOylation Of Nur77 And Functions In Breast Cancer Cells

There are such a lot members in nuclear receptor superfamily, and they join in multiple activities with large kind of down-regulators. In the progress of regulating cells, most of nuclear receptors have specific ligand-activated, except the orphan nuclear receptor subfamily. Ligands of the them are unknown, or they exercise functions without ligand-activated such as NR4A (Nuclear receptor subfamily 4 group A). Nur77 (also called NR4A1) has significant effects in serious activities, like in regulating of neurodevelopment, stimulate response and so on. The value of Nur77 gradually appears in prostatic cancer, lung cancer, bladder cancer and other cancers, proves that high expression of Nur77 consists with occurrence of cancers. Nur77 expresses much more in breast cancer cells (BCs) than normal breast cells, the influence of this phenomena has not been determined yet. Estrogen receptor a (ERa) is a primary factor in BCs and the main core of breast cancer researches. In judging the effection of Nur77 in BCs, its relationship with ERa will become the focus of the study. As Small ubiquitin-related modifer, SUMOylation is so similar to ubiquitination, such as in the choice on structure and modification site of substrates. SUMOylation can greatly affect protein intracellular localization, stability, and so on. Member of the NR4A family members Nurrl (also called NR4A2) can be modificated by SUMO. Nur77 has conservate structure and function with Nurrl, but SUMOylation of its has not been confirmed.To sum up, this study confirmed that Nur77 as a co-activator of ERa promoted growth of BCs; meanwhile, Nur77 could be modificated by SUMO, and SUMOylation inhibited its functions in BCs, the contents are as follows:1. As a substrate, Nur77 had two potential site for SUMOylation by bio informatics analysis. Its SUMOylation was demonstrated in Hela cells by immunofluorescence and immunoprecipitation experiment. Immunofluorescence revealed the colocalization of them in cell. For further confirmed, three kinds of mutant were constructed by the way of site-directed mutagenesis. Compare with wide-type, K577R and 2KR failed to covalent binding with SUMO in immunoprecipitation experiment, indicated K577 was the main modification site.2. The interaction of Nur77 and ERa was shown in Hela cells. By luciferase reporter experiment, Nur77 assumed as a co-activator which had a dose-dependent positive regulation to transcriptional activity of ERa.3. SUMOylation effected functions of Nur77, such as cell-location, transcriptional activity and promotion in breast cancer cells. That demonstrated by immunofluorescence, luciferase reporter, cell-proliferation MTT and clone formation treatment.In a word, this research contacted Nur77 to ERa, and confirmed Nur77 could be SUMOylated. Then expanded functions of Nur77, and offered a new potentiality in the therapy of breast cancer.