The Menin Tumor Suppressor Protein Is Phosphorylated in Response to DNA Damage

Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, United States of America.
PLoS ONE (Impact Factor: 3.23). 01/2011; 6(1):e16119. DOI: 10.1371/journal.pone.0016119
Source: PubMed


Multiple endocrine neoplasia type 1 (MEN1) is a heritable cancer syndrome characterized by tumors of the pituitary, pancreas and parathyroid. Menin, the product of the MEN1 gene, is a tumor suppressor protein that functions in part through the regulation of transcription mediated by interactions with chromatin modifying enzymes.
Here we show menin association with the 5' regions of DNA damage response genes increases after DNA damage and is correlated with RNA polymerase II association but not with changes in histone methylation. Furthermore, we were able to detect significant levels of menin at the 3' regions of CDKN1A and GADD45A under conditions of enhanced transcription following DNA damage. We also demonstrate that menin is specifically phosphorylated at Ser394 in response to several forms of DNA damage, Ser487 is dynamically phosphorylated and Ser543 is constitutively phosphorylated. Phosphorylation at these sites however does not influence the ability to interact with histone methyltransferase activity. In contrast, the interaction between menin and RNA polymerase II is influenced by phosphorylation, whereby a phospho-deficient mutant had a higher affinity for the elongating form of RNA polymerase compared to wild type. Additionally, a subset of MEN1-associated missense point mutants, fail to undergo DNA damage dependent phosphorylation.
Together, our findings suggest that the menin tumor suppressor protein undergoes DNA damage induced phosphorylation and participates in the DNA damage transcriptional response.

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Available from: Joshua M Francis, Dec 18, 2013
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    ABSTRACT: Most patients (70-90%) with the multiple endocrine neoplasia type 1 (MEN1) syndrome possess germline heterozygous mutations in MEN1 that predisposes to tumors of multiple endocrine and nonendocrine tissues. Some endocrine tumors of the kinds seen in MEN1 that occur sporadically in the general population also possess somatic mutations in MEN1. Interestingly, the endocrine tumors of MEN1 are recapitulated in mouse models of Men1 loss that serve as a valuable resource to understand the pathophysiology and molecular basis of tumorigenesis. Exploring these endocrine tumors in mouse models using in vivo, ex vivo and in vitro methods can help to follow the process of tumorigenesis, and can be useful for preclinical testing of therapeutics and understanding their mechanisms of action.
    Preview · Article · Nov 2014
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    ABSTRACT: In multiple endocrine neoplasia type 1 (MEN1) characterized by tumors of parathyroid, enteropancreas, and anterior pituitary, missense mutations in the MEN1 gene product, menin, occur in a subset of cases. The mutant proteins are degraded by the proteasome. However, whether their expression and activity can be restored is not known. Our objective was to functionally characterize a panel of 16 menin missense mutants, including W423R and S443Y identified in new MEN1 families, with respect to protein stability, targeting to the proteasome and restoration of expression by proteasome inhibitors and expression and function by small interfering RNA technology. Flag-tagged wild-type (WT) and missense menin mutant expression vectors were transiently transfected in human embryonic kidney (HEK293) and/or rat insulinoma (Rin-5F) cells. The majority of mutants were short-lived, whereas WT menin was stable. Proteasome inhibitors MG132 and PS-341 and inhibition of the chaperone, heat-shock protein 70 (Hsp70), or the ubiquitin ligase, COOH terminus of Hsp70-interacting protein (CHIP), by specific small interfering RNA, restored the levels of the mutants, whereas that of WT menin was largely unaffected. Inhibition of CHIP restored the ability of mutants to mediate normal functions of menin: TGF-β up-regulation of the promoters of its target genes, the cyclin-dependent kinase inhibitors p15 and p21 as well as TGF-β inhibition of cell numbers. When the levels of missense menin mutants that are targeted to the proteasome are normalized they may function similarly to WT menin. Potentially, targeting specific components of the proteasome chaperone pathway could be beneficial in treating a subset of MEN1 cases.
    No preview · Article · Nov 2011 · The Journal of Clinical Endocrinology and Metabolism
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    ABSTRACT: The aim of this study was to elucidate the expression and localization of menin, a protein encoded by the multiple endocrine neoplasia type I (MEN1) gene, in 13 human cancer cell lines. Reverse transcription-polymerase chain reaction (RT-PCR) was used to determine the expression of the menin gene. The localization of the menin protein was detected by immunofluorescence microscopy. Western blotting was used to determine the quantity of menin in the nucleus, cytosol and membrane of the cells. RT-PCR revealed that menin was expressed in all the cell lines examined in this study. Immunofluorescence microscopy revealed that menin was located primarily in the nucleus. In the GES-1 (transformed human gastric epithelium), MCF-7 (breast cancer), SGH44 (brain glioma) and HeLa (cervical cancer) cell lines, menin was also found to be localized to the membrane, cytosol and nucleus. Moreover, in SGH44 cells more menin was located in the cytosol than the nucleus. Similar findings were obtained by western blotting. In the GES-1 and MKN-28 cells undergoing octreotide treatment, cytoplasmic menin was significantly increased compared with the control groups. Therefore, we suggest that menin is expressed in a number of human cancer cell lines and that the cytosolic distribution increases when the cells undergo octreotide treatment, indicating a new role for menin.
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