title: Prohibitin 2 represents a novel nuclear AKT substrate during all-trans retinoic acid-induced differentiation of acute promyelocytic leukemia cells.
abstract: The AKT/PKB kinase is essential for cell survival, proliferation, and differentiation; however, aberrant AKT activation leads to the aggressiveness and drug resistance of many human neoplasias. In the human acute promyelocytic leukemia cell line NB4, nuclear AKT activity increases during all-trans retinoic acid (ATRA)-mediated differentiation. As nuclear AKT activity is associated with differentiation, we sought to identify the nuclear substrates of AKT that were phosphorylated after ATRA treatment. A proteomics-based search for nuclear substrates of AKT in ATRA-treated NB4 cells was undertaken by using 2D-electrophoresis/mass spectrometry (MS) in combination with an anti-AKT phospho-substrate antibody. Western blot analysis, an in vitro kinase assay, and/or site-directed mutagenesis were performed to further characterize the MS findings. MS analysis revealed prohibitin (PHB)-2, a multifunctional protein involved in cell cycle progression and the suppression of oxidative stress, to be a putative nuclear substrate of AKT. Follow-up studies confirmed that AKT phosphorylates PHB2 on Ser-91 and that forced expression of the PHB2(S91A) mutant results in a rapid loss of viability and apoptotic cell death. Activation of nuclear AKT during ATRA-mediated differentiation results in the phosphorylation of several proteins, including PHB2, which may serve to coordinate nuclear-mitochondrial events during differentiation.
authors: 2IGM-CNR, Bologna, Rizzoli Orthopedic Institute, via di Barbiano, 1/10, 40136 Bologna, Italy. william.blalock@cnr.it.
authors: Alberto
authors: A
authors: Bavelloni
authors: Manuela
authors: M
authors: Piazzi
authors: Irene
authors: I
authors: Faenza
authors: Mirco
authors: M
authors: Raffini
authors: Antonietta
authors: A
authors: D'Angelo
authors: Luca
authors: L
authors: Cattini
authors: Lucio
authors: L
authors: Cocco
authors: William L
authors: WL
authors: Blalock
mesh_terms: D017209:Apoptosis; D002453:Cell Cycle; D002454:Cell Differentiation; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002470:Cell Survival; D057809:HEK293 Cells; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D018384:Oxidative Stress; D010766:Phosphorylation; D000091402:Prohibitins; D040901:Proteomics; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism*; D015398:Signal Transduction; D014212:Tretinoin / Q000378:metabolism*
chemical_list: C000717328:PHB protein, human; C000717331:PHB2 protein, human; D000091402:Prohibitins; D012097:Repressor Proteins; D014212:Tretinoin; D051057:Proto-Oncogene Proteins c-akt
keywords: SRp20/SRSF3; mass spectrometry; phosphorylation; proteomics; signal transduction