Polyamines modulate the subcellular localization of RNA-binding protein HuR through AMP-activated protein kinase-regulated phosphorylation and acetylation of importin alpha1.
Polyamines are required for maintenance of intestinal epithelial integrity, and a decrease in cellular polyamines increases the cytoplasmic levels of RNA-binding protein HuR stabilizing p53 and nucleophosmin mRNAs, thus inhibiting IEC (intestinal epithelial cell) proliferation. The AMPK (AMP-activated protein kinase), an enzyme involved in responding to metabolic stress, was recently found to be implicated in regulating the nuclear import of HuR. Here, we provide evidence showing that polyamines modulate subcellular localization of HuR through AMPK-regulated phosphorylation and acetylation of Impalpha1 (importin alpha1) in IECs. Decreased levels of cellular polyamines as a result of inhibiting ODC (ornithine decarboxylase) with DFMO (D,L-alpha-difluoromethylornithine) repressed AMPK activity and reduced Impalpha1 levels, whereas increased levels of polyamines as a result of ODC overexpression induced both AMPK and Impalpha1 levels. AMPK activation by overexpression of the AMPK gene increased Impalpha1 but reduced the cytoplasmic levels of HuR in control and polyamine-deficient cells. IECs overexpressing wild-type Impalpha1 exhibited a decrease in cytoplasmic HuR abundance, while cells overexpressing Impalpha1 proteins bearing K22R (lacking acetylation site), S105A (lacking phosphorylation site) or K22R/S105A (lacking both sites) mutations displayed increased levels of cytoplasmic HuR. Ectopic expression of these Impalpha1 mutants also prevented the increased levels of cytoplasmic HuR following polyamine depletion. These results indicate that polyamine-mediated AMPK activation triggers HuR nuclear import through phosphorylation and acetylation of Impalpha1 in IECs and that polyamine depletion increases cytoplasmic levels of HuR as a result of inactivation of the AMPK-driven Impalpha1 pathway.
Zou
Tongtong
T
Cell Biology Group, Department of Surgery, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
Liu
Lan
L
Rao
Jaladanki N
JN
Marasa
Bernard S
BS
Chen
Jie
J
Xiao
Lan
L
Zhou
Huiping
H
Gorospe
Myriam
M
Wang
Jian-Ying
JY
D055372:AMP-Activated Protein Kinases; D000107:Acetylation; D000818:Animals; D000954:Antigens, Surface / Q000032:analysis* / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D006801:Humans; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000378:metabolism*; D010766:Phosphorylation; D011073:Polyamines / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000032:analysis* / Q000378:metabolism*; D051381:Rats; D015398:Signal Transduction; D028901:alpha Karyopherins / Q000378:metabolism*
D000954:Antigens, Surface; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; C098770:ELAVL1 protein, human; D009097:Multienzyme Complexes; D011073:Polyamines; D016601:RNA-Binding Proteins; D028901:alpha Karyopherins; C087978:karyopherin alpha 2; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases; C526180:PRKAA1 protein, human