Identification of multiple nuclear export sequences in Fanconi anemia group A protein that contribute to CRM1-dependent nuclear export

Department of Medical Oncology, VU University Medical Center, Amsterdam, The Netherlands.
Human Molecular Genetics (Impact Factor: 6.68). 06/2005; 14(10):1271-81. DOI: 10.1093/hmg/ddi138
Source: PubMed

ABSTRACT The Fanconi anemia (FA) pathway plays an important role in maintaining genomic stability, and defects in this pathway cause cancer susceptibility. The FA proteins have been found to function primarily in a nuclear complex, although a cytoplasmic localization and function for several FA proteins has also been reported. In this study, we investigated the possibility that FANCA, FANCC and FANCG are subjected to active export out of the nucleus. After treatment with leptomycin B, a specific inhibitor of CRM1-mediated nuclear export, the accumulation of epitope-tagged FANCA in the nucleus increased, whereas FANCC was affected to a lesser extent and FANCG showed no response. CRM1-mediated export of FANCA was further confirmed using CRM1 cotransfection, which led to a dramatic relocalization of FANCA to the cytoplasm. Five functional leucine-rich nuclear export sequences (NESs) distributed throughout the FANCA sequence were identified and characterized using an in vivo export assay. Simultaneous inactivation of three of these NESs resulted in a discrete but reproducible increase of FANCA nuclear accumulation. However, these NES mutations did not affect the ability of FANCA to complement the mitomycin C or cisplatin sensitivity of FA-A lymphoblasts. Surprisingly, mutations in the other two NESs resulted in an almost complete relocation of the protein to cytoplasm, suggesting that these motifs overlap with domains that are crucial for nuclear import. Taken together, these findings indicate that FANCA can be actively exported out of the nucleus by CRM1, revealing a new mechanism to regulate the function of the FA protein complex.

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Available from: Frank Kruyt, Jan 08, 2015
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    • "Comparable mutations of leucine residues to alanines have frequently been used for the inactivation of NES sites (Ferrer et al., 2005; Jensik et al., 2004; Wen et al., 1995). NES sites are frequently and preferentially located within α-helical structures, and the most C-terminal leucine tends to be exposed and available for interactions (La Cour et al., 2004). "
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    Neurobiology of Disease 07/2013; DOI:10.1016/j.nbd.2009.07.020 · 5.20 Impact Factor
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    • "In some instances the loss of FANCA, FANCC, FANCE or FANCF results in the formation of smaller sub-complexes, which are incompetent at monoubiquitinating FANCD2 [9] [10] [11]. Furthermore, there is some evidence indicating that the nuclear import of some FA proteins (FANCA and FANCC) is regulated [12] [13]. Since the assembly of the complete core complex requires the availability of all the constituents in the nucleus, it is conceivable that the pathway may be regulated at the point of assembly (Fig. 2B). "
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    ABSTRACT: There has been a recent profusion of reviews on Fanconi anemia (FA), which will give readers a comprehensive outline of the field R.D. Kennedy, A.D. D'Andrea, The Fanconi anemia/BRCA pathway: new faces in the crowd, Genes Dev. 19 (2005) 2925-2940; L.J. Niedernhofer, A.S. Lalai, J.H. Hoeijmakers, Fanconi anemia (cross)linked to DNA repair, Cell 123 (2005) 1191-1198; H. Joenje, K.J. Patel, The emerging genetic and molecular basis of Fanconi anaemia, Nat. Rev. Genet. 2 (2001) 446-457. Here, we will focus on key areas that place the FA proteins in the context of DNA repair during replication. In addition, where possible we will put forward propositions that in our opinion need addressing, and where possible provide models that can be tested.
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