Sphingosine interaction with acidic leucine-rich nuclear phosphoprotein-32A (ANP32A) regulates PP2A activity and cyclooxygenase (COX)-2 expression in human endothelial cells.
Sphingolipid metabolites regulate cell fate by acting on specific cellular targets. Although the influence of sphingolipids in cellular signaling has been well recognized, the exact molecular targets and how these targets influence cellular signaling mechanisms remain poorly understood. Toward this goal, we used affinity chromatography coupled with proteomics technology and identified acidic leucine-rich nuclear phosphoprotein-32A (ANP32A), an inhibitor of protein phosphatase 2A (PP2A) as a direct target of sphingosine, N,N'-dimethyl sphingosine (DMS) and phytosphingosine but not dihydrosphingosine or sphingosine 1-phosphate. Treatment of human umbilical vein endothelial cells (HUVEC) with DMS, which is not phosphorylated by sphingosine kinases, led to the activation of PP2A activity. Suppression of ANP32A with siRNA enhanced basal and DMS-activated PP2A activity suggesting that the sphingoid base binds to and relieves the inhibitory action of ANP32A on the PP2A complex. Indeed, DMS relieved the ANP32A-mediated inhibition of PP2A enzyme complex in vitro. Interestingly, DMS treatment induced the p38 stress-activated protein kinase (SAPK) and expression of cyclooxygenase (COX)-2 transcript and protein. Knockdown of ANP32A expression further induced p38 SAPK and COX-2. These data identify ANP32A as a novel molecular target of sphingoid bases that regulates cellular signaling events and inflammatory gene expression.
Habrukowich
Cheryl
C
Center for Vascular Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3501.
Han
David K
DK
Center for Vascular Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3501.
Le
Andrew
A
Calhoun Cardiology Center, Department of Cell Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3501.
Rezaul
Karim
K
Center for Vascular Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3501.
Pan
Wei
W
Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021.
Ghosh
Mallika
M
Center for Vascular Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3501.
Li
Zaiguo
Z
Department of Chemistry and Biochemistry, Queens College, City University of New York, New York, New York 11367.
Dodge-Kafka
Kimberly
K
Calhoun Cardiology Center, Department of Cell Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3501.
Jiang
Xuejun
X
Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021.
Bittman
Robert
R
Department of Chemistry and Biochemistry, Queens College, City University of New York, New York, New York 11367.
Hla
Timothy
T
Center for Vascular Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3501; Center for Vascular Biology, Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, Cornell University, New York, New York 10065. Electronic address: tih2002@med.cornell.edu.
D002460:Cell Line; D051546:Cyclooxygenase 2 / Q000096:biosynthesis* / Q000235:genetics; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D048670:MAP Kinase Kinase 4 / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins; D054648:Protein Phosphatase 2 / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D016601:RNA-Binding Proteins; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology
C087425:ANP32A protein, human; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D048670:MAP Kinase Kinase 4; C517189:PPP2CA protein, human; D054648:Protein Phosphatase 2; C061800:N,N-dimethylsphingosine; D013110:Sphingosine