A nuclear export signal and oxidative stress regulate ShcD subcellular localisation: A potential role for ShcD in the nucleus

• Published in: Cellular signalling 2014-01, Vol.26 (1), p.32-40
• Main Authors: Ahmed, Samrein B.M., Prigent, Sally A.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Cell Nucleus - drug effects; Cell Nucleus - metabolism; Deoxyribonucleic acid; DNA - metabolism; Exports; Gene Expression Regulation - drug effects; HEK293 Cells; Histones - metabolism; Humans; Hydrogen Peroxide - pharmacology; International trade; Kinases; Luciferases - metabolism; Molecular Sequence Data; Nuclear export signal; Nuclear Export Signals; Nuclei; Oxidative stress; Oxidative Stress - drug effects; Protein Structure, Tertiary; Protein Transport - drug effects; Proteins; Shc; Shc Signaling Adaptor Proteins - chemistry; Shc Signaling Adaptor Proteins - metabolism; Signalling; Signalling adaptor; Stresses; Structure-Activity Relationship; Subcellular Fractions - drug effects; Subcellular Fractions - metabolism; Time-Lapse Imaging
• ISSN: 0898-6568; 1873-3913
• DOI: 10.1016/j.cellsig.2013.09.003

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 83LCTLIPRM90 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. •A nuclear export sequence, 83LCTLIPRM90 is present in the CH2 domain of p69ShcD•This sequence is required for cytoplasmic localisation of p69ShcD•Shorter isoforms; p59ShcD and p49ShcD are present in the nucleus•A proportion of p69ShcD translocates to the nucleus upon oxidative stress•Nuclear ShcD may have a role in regulating transcription