Journal
MOLECULAR AND CELLULAR BIOLOGY
Volume 23, Issue 2, Pages 493-509Publisher
AMER SOC MICROBIOLOGY
DOI: 10.1128/MCB.23.2.493-509.2003
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Funding
- NATIONAL HUMAN GENOME RESEARCH INSTITUTE [ZIAHG000029, Z01HG000029] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES [Z01DK043009, ZIEDK043006, ZIEDK043009, Z01DK043006] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE ON DEAFNESS AND OTHER COMMUNICATION DISORDERS [Z01DC000062] Funding Source: NIH RePORTER
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Menin is a 70-kDa protein encoded by MEN1, the tumor suppressor gene disrupted in multiple endocrine neoplasia type 1. In a yeast two-hybrid system based on reconstitution of Ras signaling, menin was found to interact with the 32-kDa subunit (RPA2) of replication protein A (RPA), a heterotrimeric protein required for DNA replication, recombination, and repair. The menin-RPA2 interaction was confirmed in a conventional yeast two-hybrid system and by direct interaction between purified proteins. Menin-RPA2 binding was inhibited by a number of menin missense mutations found in individuals with multiple endocrine neoplasia type 1, and the interacting regions were mapped to the N-terminal portion of menin and amino acids 43 to 171 of RPA2. This region of RPA2 contains a weak single-stranded DNA-binding domain, but menin had no detectable effect on RPA-DNA binding in vitro. Menin bound preferentially in vitro to free RPA2 rather than the RPA heterotrimer or a subcomplex consisting of RPA2 bound to the 14-kDa subunit (RPA3). However, the 70-kDa subunit (RPA1) was coprecipitated from HeLa cell extracts along with RPA2 by menin-specific antibodies, suggesting that menin binds to the RPA heterotrimer or a novel RPA1-RPA2-containing complex in vivo. This finding was consistent with the extensive overlap in the nuclear localization patterns of endogenous menin, RPA2, and RPA1 observed by immunofluorescence.
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