Nuclear and nucleolar localization signals and their targeting function in phosphatidylinositol 4-kinase PI4K230

Journal Article · Fri Aug 01 04:00:00 EDT 2008 · Experimental Cell Research

DOI:https://doi.org/10.1016/j.yexcr.2008.05.006· OSTI ID:21128143

Kakuk, Annamaria ^[1]; Friedlaender, Elza; Vereb, Gyoergy; Lisboa, Duarte ^[2]; Bagossi, Peter ^[3]; Toth, Gabor ^[4]; Gergely, Pal ^[1]; Vereb, Gyoergy ^[1]

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1. Department of Medical Chemistry, Medical and Health Science Center, University of Debrecen, H-4032 Debrecen, Nagyerdei krt. 98 (Hungary)
2. Department of Biophysics and Cell Biology, Medical and Health Science Center, University of Debrecen, H-4032 Debrecen, Nagyerdei krt. 98 (Hungary)
3. Department of Biochemistry and Molecular Biology, Medical and Health Science Center, University of Debrecen, H-4032 Debrecen, Nagyerdei krt. 98 (Hungary)
4. Institute for Medical Chemistry, Szeged University of Sciences, 6720 Szeged, Dom ter 8 (Hungary)

PI4K230, an isoform of phosphatidylinositol 4-kinase, known primarily as a cytoplasmic membrane-bound enzyme, was detected recently also in the nucleolus of several cells. Here we provide mechanistic insight on the targeting function of its putative nuclear localization signal (NLS) sequences using molecular modeling, digitonin-permeabilized HeLa cells and binding to various importins. The synthetic sequence {sup 916}NFNHIHKRIRRVADKYLSG{sup 934} comprising a putative monopartite NLS (NLS1), targeted covalently bound fluorescent BSA to the nucleoplasm via classical importin {alpha}/{beta} mechanism employing importins {alpha}1 and {alpha}3 but not {alpha}5. This transport was inhibited by wheat germ agglutinin and GTP{gamma}S. The sequence {sup 1414}SKKTNRGSQLHKYYMKRRTL{sup 1433}, a putative bipartite NLS (NLS2) proved ineffective in nuclear targeting if conjugated to fluorescently labeled BSA. Nonetheless, NLS2 or either of its basic clusters directed to the nucleolus soybean trypsin inhibitor that can pass the nuclear pore complex passively; moreover, an expressed 58 kDa fragment of PI4K230 (AA1166-1667) comprising NLS2 was also imported into the nucleus by import factors of reticulocyte lysate or by importin {alpha}1/{beta} or {alpha}3/{beta} complexes and localized to the nucleolus. We conclude that the putative bipartite NLS itself is a nucleolar targeting signal, and for nuclear import PI4K230 requires a larger sequence around it or, alternatively, the monopartite NLS.

• OSTI ID: 21128143
• Journal Information: Experimental Cell Research, Journal Name: Experimental Cell Research Journal Issue: 13 Vol. 314; ISSN 0014-4827; ISSN ECREAL
• Country of Publication: United States
• Language: English

Related Subjects

60 APPLIED LIFE SCIENCES
AGGLUTININS
CELL NUCLEI
HELA CELLS
SOYBEANS
TRYPSIN
WHEAT