title: Proteomic analysis of NME1/NDPK A null mouse liver: evidence for a post-translational regulation of annexin IV and EF-1Bα.
abstract: NME/NDPK family proteins are involved in the control of intracellular nucleotide homeostasis as well as in both physiological and pathological cellular processes, such as proliferation, differentiation, development, apoptosis, and metastasis dissemination, through mechanisms still largely unknown. One family member, NME1/NDPK-A, is a metastasis suppressor, yet the primary physiological functions of this protein are still missing. The purpose of this study was to identify new NME1/NDPK-A-dependent biological functions and pathways regulated by this gene in the liver. We analyzed the proteomes of wild-type and transgenic NME1-null mouse livers by combining two-dimensional gel electrophoresis and mass spectrometry (matrix-assisted laser desorption/ionization time of flight and liquid chromatography-tandem mass spectrometry). We found that the levels of three proteins, namely, phenylalanine hydroxylase, annexin IV, and elongation factor 1 Bα (EF-1Bα), were strongly reduced in the cytosolic fraction of NME1(-/-) mouse livers when compared to the wild type. This was confirmed by immunoblotting analysis. No concomitant reduction in the corresponding messenger RNAs or of total protein level was observed, however, suggesting that NME1 controls annexin IV and EF-1Bα amounts by post-translational mechanisms. NME1 deletion induced a change in the subcellular location of annexin IV in hepatocytes resulting in enrichment of this protein at the plasma membrane. We also observed a redistribution of EF-1Bα in NME1(-/-) hepatocytes to an intracytoplasmic compartment that colocalized with a marker of the reticulum endoplasmic. Finally, we found reduced expression of annexin IV coincident with decreased NME1 expression in a panel of different carcinoma cell lines. Taken together, our data suggest for the first time that NME1 might regulate the subcellular trafficking of annexin IV and EF-1Bα. The potential role of these proteins in metastatic dissemination is discussed.
authors.affiliation: Service de Biochimie A, Hôpital Saint-Antoine, AP-HP, 75012, Paris, France.
authors.forename: Arnaud
authors.initials: A
authors.lastname: Bruneel
authors.forename: Dominique
authors.initials: D
authors.lastname: Wendum
authors.forename: Valérie
authors.initials: V
authors.lastname: Labas
authors.forename: Odile
authors.initials: O
authors.lastname: Mulner-Lorillon
authors.forename: Joelle
authors.initials: J
authors.lastname: Vinh
authors.forename: Nelly
authors.initials: N
authors.lastname: Bosselut
authors.forename: Eric
authors.initials: E
authors.lastname: Ballot
authors.forename: Bruno
authors.initials: B
authors.lastname: Baudin
authors.forename: Chantal
authors.initials: C
authors.lastname: Housset
authors.forename: Sandrine
authors.initials: S
authors.lastname: Dabernat
authors.forename: Marie-Lise
authors.initials: ML
authors.lastname: Lacombe
authors.forename: Mathieu
authors.initials: M
authors.lastname: Boissan
mesh_terms: D000818:Animals; D017314:Annexin A4 / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D003600:Cytosol / Q000201:enzymology / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004721:Endoplasmic Reticulum / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000201:enzymology* / Q000378:metabolism; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D054778:NM23 Nucleoside Diphosphate Kinases / Q000235:genetics / Q000502:physiology*; D020648:Peptide Elongation Factor 1 / Q000235:genetics* / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D040901:Proteomics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry
chemical_list: D017314:Annexin A4; D054778:NM23 Nucleoside Diphosphate Kinases; D020648:Peptide Elongation Factor 1; C517939:Nme1 protein, mouse