期刊
MOLECULAR BIOLOGY OF THE CELL
卷 14, 期 3, 页码 1221-1239出版社
AMER SOC CELL BIOLOGY
DOI: 10.1091/mbc.E02-03-0170
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- NCI NIH HHS [CA-71514] Funding Source: Medline
- NIDDK NIH HHS [R56 DK042799, DK-42799, R01 DK042799] Funding Source: Medline
The transcriptionally regulated serum and glucocorticoid inducible protein kinase (Sgk) is localized to the nucleus in a serum-dependent manner, and a yeast two-hybrid genetic screen uncovered a specific interaction between Sgk and the importin-alpha nuclear import receptor. In vitro GST pull down assays demonstrated a strong and direct association of importin-alpha with endogenous Sgk and exogenously expressed HA-tagged Sgk, whereas both components coimmunoprecipitate and colocalize to the nucleus after serum stimulation. Consistent with an active mechanism of nuclear localization, the nuclear import of HA-Sgk in permeabilized cells required ATP, cytoplasm, and a functional nuclear pore complex. Ectopic addition of a 107 amino acid carboxyterminal fragment of importin-alpha, which contains the Sgk binding region, competitively inhibited the ability of endogenous importin-alpha to import Sgk into nuclei in vitro. Mutagenesis of lysines by alanine substitution defined a KKAILKKKEEK sequence within the central domain of Sgk between amino acids 131-141 that functions as a nuclear localization signal (NLS) required for the in vitro interaction with importin-alpha and for nuclear import of full-length Sgk in cultured cells. The serum-induced nuclear import of Sgk requires the NLS-dependent recognition of Sgk by importin-alpha as well as the PI3-kinase- dependent phosphorylation of Sgk. Our results define a new role importin-alpha in the stimulus-dependent control of signal transduction by nuclear localized protein kinases.
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