title: A nuclear export signal and oxidative stress regulate ShcD subcellular localisation: a potential role for ShcD in the nucleus.
abstract: Tumour cells alter their gene expression profile to acquire a more invasive and resistant phenotype. Overexpression of the signalling adaptor protein ShcD in melanoma was found to be a prerequisite for melanoma migration and invasion. In common with other Shc proteins, ShcD has been shown to be involved in coupling receptor tyrosine kinases to the Ras-mitogen activated protein kinase signalling pathway, and to have a predominant cytoplasmic distribution. Here we report that ShcD can exist within the nucleus, and show that its CH2 domain has a critical role in nuclear export of ShcD. Analysis of GFP-tagged ShcD mutants containing deletions or amino acid substitutions within the CH2 domain revealed (83)LCTLIPRM(90) as a functional nuclear export signal. We have further demonstrated that ShcD accumulates in the nucleus upon hydrogen peroxide treatment in FLAG-ShcD expressing HEK293 cells, as well as 518.A2 melanoma cells. Cross linking experiments showed that a proportion of ShcD is associated with DNA. Moreover we have shown that ShcD fused to the GAL4 DNA binding domain can drive transcription of a GAL4 site-driven luciferase reporter, suggesting a role for ShcD in regulating gene transcription. We suggest that ShcD nuclear translocation might provide melanoma cells with a mechanism that enables them to resist DNA damage due to oxidative stress.
authors.affiliation: Department of Biochemistry, University of Leicester, Lancaster Road, Leicester, LE1 9HN, United Kingdom.
authors.forename: Samrein B M
authors.initials: SB
authors.lastname: Ahmed
authors.forename: Sally A
authors.initials: SA
authors.lastname: Prigent
mesh_terms: D000595:Amino Acid Sequence; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D057809:HEK293 Cells; D006657:Histones / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008156:Luciferases / Q000378:metabolism; D008969:Molecular Sequence Data; D049790:Nuclear Export Signals; D018384:Oxidative Stress / Q000187:drug effects; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000187:drug effects; D055765:Shc Signaling Adaptor Proteins / Q000737:chemistry / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism; D059008:Time-Lapse Imaging
chemical_list: D006657:Histones; D049790:Nuclear Export Signals; C519335:SHC4 protein, human; D055765:Shc Signaling Adaptor Proteins; D004247:DNA; D006861:Hydrogen Peroxide; D008156:Luciferases
keywords: Nuclear export signal; Oxidative stress; Shc; Signalling adaptor