Correlation of mutant menin stability with clinical expression of multiple endocrine neoplasia type 1 and its incomplete forms

Abstract

Germline mutations of the tumor suppressor gene MEN1 are found not only in typical multiple endocrine neoplasia type 1 (MEN1) but also in its incomplete forms such as familial isolated hyperparathyroidism (FIHP) and apparently sporadic parathyroid tumor (ASPT). No definitive genotype-phenotype correlation has been established between these clinical forms and MEN1 gene mutations. We previously demonstrated that mutant menin proteins associated with MEN1 are rapidly degraded by the ubiquitin-proteasome pathway. To examine whether the intracellular stability of mutant menin is correlated with clinical phenotypes, we developed a method of evaluating menin stability and examined 20 mutants associated with typical MEN1 (17 missense, two in-frame deletion, one nonsense) and 21 mutants associated with FIHP or ASPT (19 missense, two in-frame deletion). All tested mutants associated with typical MEN1 showed reduced stability. Some missense and in-frame deletion mutants (G28A, R171W, T197I, E255K, E274A, Y353del and E366D) associated with FIHP or ASPT were almost as stable as or only slightly less stable than wild-type menin, while others were as unstable as those associated with typical MEN1. Some stable mutants exhibited substantial biological activities when tested by JunD-dependent transactivation assay. These findings suggest that certain missense and in-frame mutations are fairly stable and retain intrinsic biological activity, and might be specifically associated with incomplete clinical phenotypes. The menin stability test will provide useful information for the management of patients carrying germline MEN1 mutations especially when they have missense or in-frame variants of ambiguous clinical significance. (Cancer Sci 2011; 102: 2097-2102 M enin is a tumor suppressor protein encoded by MEN1, a gene responsible for multiple endocrine neoplasia type 1 (MEN1), a familial cancer syndrome typically characterized by the development of multiple tumors in the pituitary, parathyroid and enteropancreatic endocrine tissues.(1,2) Menin is a nuclear protein having C-terminal nuclear localizing signal sequences, (3) and exhibits modulatory activity on gene expression such as repression of JunD-dependent transcription. (4) Diverse roles have been implicated for menin, including cell cycle control, cell differentiation and DNA repair, which are likely to be conferred by interaction with various menin-binding proteins.(2) Menin is considered to be a scaffold protein tethering such proteins to specific gene loci.(5) Although the mechanism of how menin is involved in gene regulation has been elucidated to some extent, the basis for tissue-specific tumorigenesis in MEN1 remains unknown.Heterozygous germline mutations of the MEN1 gene are found in 70-90% of patients clinically diagnosed as MEN1. (6) More than 500 different loss-of-function mutations have been identified, which are distributed throughout the gene with no apparent hot spot.(2,6,7) Germline MEN1 mutations have also been found in a subset of familial isolated hyperparathy...

Application of an intracellular stability test of a novel missense menin mutant to the diagnosis of multiple endocrine neoplasia type 1

“…The intracellular stability of missense menin variants was evaluated using a quantitative fluorescent immunohistochemical method as described previously [15,16]. Briefly, WI38VA13 cells were transfected with a bicistronic plasmid expressing N-terminal FLAG-tagged and Myc-tagged proteins: one protein was wild type menin, which served as an internal control for transfection efficiency, and the other was the variant menin to be tested.…”

“…We are also aware that the stability of menin missense mutants is highly variable and that some mutants associated with typical MEN1 are comparatively stable [16,19]. Therefore, the pathogenicity of a missense mutation giving rise to a stable mutant menin should be interpreted cautiously.…”

“…and stability of missense menin tested in various nonendocrine as well as in endocrine cells [15,16]. We are also aware that the stability of menin missense mutants is highly variable and that some mutants associated with typical MEN1 are comparatively stable [16,19].…”

“…We previously reported that missense mutant menin proteins associated with MEN1 are unstable and rapidly degraded through ubiquitin-proteasome pathway [15]. More detailed analysis by a newly developed fluorescence immunocytochemical method revealed that the stability of missense and in-frame deletion mutants varies widely but that unstable mutants were found only in MEN1 and related disorders and not in normal polymorphisms [16]. We recently encountered a suspected MEN1 patient with a previously unreported missense mutation in the MEN1 gene.…”

A new double substitution mutation in the MEN1 gene: a limited penetrance and a specific phenotype

“…7 Recently, authors argued that the stability of the mutant menin could correlate with the clinical expression of MEN1. 8 The MEN1 mutations associated with FIHP were as stable or as slightly less stable than the wild-type menin and showed normal interaction with JunD, one of the interacting proteins of menin. 8 A majority of MEN1 gene mutation carriers (up to 99%) will express at least one manifestation of the disease before the age of 50.…”