| First Author | Kirby TW | Year | 2017 |
| Journal | Nucleic Acids Res | Volume | 45 |
| Issue | 4 | Pages | 1958-1970 |
| PubMed ID | 27956495 | Mgi Jnum | J:247131 |
| Mgi Id | MGI:5923098 | Doi | 10.1093/nar/gkw1257 |
| Citation | Kirby TW, et al. (2017) DNA polymerase beta contains a functional nuclear localization signal at its N-terminus. Nucleic Acids Res 45(4):1958-1970 |
| abstractText | DNA polymerase beta (pol beta) requires nuclear localization to fulfil its DNA repair function. Although its small size has been interpreted to imply the absence of a need for active nuclear import, sequence and structural analysis suggests that a monopartite nuclear localization signal (NLS) may reside in the N-terminal lyase domain. Binding of this domain to Importin alpha1 (Impalpha1) was confirmed by gel filtration and NMR studies. Affinity was quantified by fluorescence polarization analysis of a fluorescein-tagged peptide corresponding to pol beta residues 2-13. These studies indicate high affinity binding, characterized by a low micromolar Kd, that is selective for the murine Importin alpha1 (mImpalpha1) minor site, with the Kd strengthening to approximately 140 nM for the full lyase domain (residues 2-87). A further reduction in Kd obtains in binding studies with human Importin alpha5 (hImpalpha5), which in some cases has been demonstrated to bind small domains connected to the NLS. The role of this NLS was confirmed by fluorescent imaging of wild-type and NLS-mutated pol beta(R4S,K5S) in mouse embryonic fibroblasts lacking endogenous pol beta. Together these data demonstrate that pol beta contains a specific NLS sequence in the N-terminal lyase domain that promotes transport of the protein independent of its interaction partners. Active nuclear uptake allows development of a nuclear/cytosolic concentration gradient against a background of passive diffusion. |
