Proteomic analysis of macrophages stimulated by lipopolysaccharide: Lipopolysaccharide inhibits the cleavage of nucleophosmin.
Lipopolysaccharide (LPS) is a complex glycolipid composed of a hydrophilic polysaccharide and a hydrophobic domain that is responsible for the biological activity of LPS. There are many reports about LPS stimulation, and many activated proteins have been detected after LPS stimulation in various cell types. Furthermore, most of the LPS signaling pathways are clear. However, we were interested in examining the changes of LPS-induced total cytosolic proteins expression and the LPS signaling pathway by the proteomics technique during LPS-induced macrophage activation. Our study employed two-dimensional gel electrophoresis and mass spectrometry to analyze the proteins involved in LPS-induced activation in RAW 264.7 cells. We found 11 protein spots whose expression was different between untreated cells and LPS-treated cells. Ten protein spots were identified, seven of which, tubulin beta-4 chain (49.6 kDa, pI 4.78), nucleophosmin (32.6 kDa, pI 4.62, two spots), 40S ribosomal protein SA (P40) (32.7 kDa, pI 4.74), transforming protein RhoA (21.8 kDa, pI 5.83), nucleolin (76.6 kDa, pI 4.69), and T-complex protein 1 zeta subunit (58 kDa, pI 6.63) were down-regulated, and three of which, nucleophosmin (32.6 kDa, pI 4.62, two spots) and proteosome subunit alpha type-1 (29.5 kDa, pI 6.00), were up-regulated. The suppression of the proteolytic degradation of nucleophosmin was associated with LPS-induced RAW 264.7 cell activation. Cleaved caspase-3 decreased, thus it might be involved in proteolysis of nucleophosmin in LPS-induced macrophage activation. Our study also demonstrated that there was no change of the expression of nucleophosmin at the mRNA level.
Zhang
Xiulian
X
Department of Biochemistry and Biomolecular Recognition, Yamaguchi University School of Medicine, Ube, Japan.
Kuramitsu
Yasuhiro
Y
Fujimoto
Masanori
M
Hayashi
Eiko
E
Yuan
Xiaoqin
X
Nakamura
Kazuyuki
K
D000818:Animals; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000096:biosynthesis; D002460:Cell Line; D002853:Chromatography, Liquid; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D008070:Lipopolysaccharides / Q000276:immunology*; D008262:Macrophage Activation; D008264:Macrophages / Q000378:metabolism*; D013058:Mass Spectrometry; D051379:Mice; D009687:Nuclear Proteins / Q000187:drug effects / Q000378:metabolism*; D000090243:Nucleophosmin; D040901:Proteomics / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D008070:Lipopolysaccharides; D009687:Nuclear Proteins; D000090243:Nucleophosmin; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases