title: BSAP (Pax5)-importin alpha 1 (Rch1) interaction identifies a nuclear localization sequence.
abstract: BSAP (Pax5) is an essential transcription factor for early B cell and central nervous system development. In later B cell development, BSAP has been implicated in the regulation of 3' Ig enhancers and a number of B cell-specific genes. Previous studies have suggested a role for BSAP-interacting proteins in the regulation of the function of BSAP. Using the yeast two-hybrid system, we identified importin alpha1 (Rch1) as a BSAP-interacting protein. Importin alpha proteins have been shown to escort proteins into the nucleus through interaction with a nuclear localization signal (NLS), composed of short stretches of basic amino acids. A predicted NLS in BSAP (NKRKRDE, located at amino acids 195-201 in the central domain) was confirmed to be essential for interaction with importin alpha1 by the yeast two-hybrid assay. Physical interaction between BSAP and importin alpha1 was detected in vitro by a glutathione S-transferase (GST) pulldown assay. The NLS sequence in BSAP conferred nuclear localization to green fluorescent protein (GFP)-BSAP fusion proteins. Although the N-terminal paired (DNA-binding) domain of BSAP also conferred nuclear localization when coupled to green fluorescent protein, this domain did not bind to importin alpha1 in the yeast two-hybrid assay. The NLS sequence in the central domain of BSAP binds to the C-terminal 98-amino acid fragment of importin alpha1.
authors.affiliation: Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
authors.forename: C R
authors.initials: CR
authors.lastname: Kovac
authors.forename: A
authors.initials: A
authors.lastname: Emelyanov
authors.forename: M
authors.initials: M
authors.lastname: Singh
authors.forename: N
authors.initials: N
authors.lastname: Ashouian
authors.forename: B K
authors.initials: BK
authors.lastname: Birshtein
mesh_terms: D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000378:metabolism*; D051757:PAX5 Transcription Factor; D011485:Protein Binding; D014157:Transcription Factors; D028901:alpha Karyopherins
chemical_list: D002352:Carrier Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D051757:PAX5 Transcription Factor; C497281:PAX5 protein, human; D014157:Transcription Factors; D028901:alpha Karyopherins; C087978:karyopherin alpha 2; D005982:Glutathione Transferase