Mass spectroscopic phosphoprotein mapping of Ral binding protein 1 (RalBP1/Rip1/RLIP76).
RalBP1, a multifunctional protein implicated in cancer cell proliferation, radiation and chemoresistance, and ligand dependent receptor internalization, is upregulated in bladder cancer and is a downstream effector of RalB, a GTPase associated with metastasis. RalBP1 can be regulated by phosphorylation by protein kinase C (PKC). No studies have comprehensively mapped RalBP1 phosphorylation sites or whether RalB affects these. We identified 14 phosphorylation sites of RalBP1 in human bladder carcinoma UMUC-3 and embryonic kidney derived 293T cells. The phosphorylated residues are concentrated at the N-terminus. Ten of the first 100 amino acids of the primary structure were phosphorylated. Nine were serine residues, and one a threonine. We evaluated the effect of RalB overexpression on RalBP1 phosphorylation and found the largest change in phosphorylation status at S463 and S645. Further characterization of these sites will provide novel insights on RalBP1 biology, its functional relationship to RalB and possible avenues for therapeutic intervention.
Herlevsen
Mikael C
MC
Department of Molecular Physiology and Biological Physics, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA.
Theodorescu
Dan
D
Department of Molecular Physiology and Biological Physics, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA. Electronic address: Dt9d@virginia.edu.
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry*; D000595:Amino Acid Sequence; D020690:GTPase-Activating Proteins / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry; D013912:Threonine / Q000737:chemistry; D001749:Urinary Bladder Neoplasms / Q000378:metabolism; D020725:ral GTP-Binding Proteins / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; D020690:GTPase-Activating Proteins; D010750:Phosphoproteins; C095637:RALBP1 protein, human; D013912:Threonine; D012694:Serine; D011493:Protein Kinase C; D020725:ral GTP-Binding Proteins
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