title: The cell senescence inducing gene product MORF4 is regulated by degradation via the ubiquitin/proteasome pathway.
abstract: After undergoing several rounds of divisions normal human fibroblasts enter a terminally non-dividing state referred to as cellular or replicative senescence. We cloned MORF4 (mortality factor on human chromosome 4), as a cellular senescence inducing gene that caused immortal cells assigned to complementation group B for indefinite division to stop dividing. To facilitate analyses of this gene, which is toxic to cells at low levels, we obtained stable clones of HeLa cells expressing a tetracycline-induced MORF4 construct that could be induced by doxycycline in a dose-dependent manner. MORF4 induction resulted in reduced colony formation after 14 days of culture, as previously observed. We determined that MORF4 protein was unstable and that addition of the proteasome inhibitor MG132 resulted in the accumulation of the protein. Following removal of MG132 the protein was rapidly degraded. Subcellular fractionation following MG132 treatment demonstrated that the protein accumulates primarily in the cytoplasm with some amounts present in the nucleus. It is therefore possible that MORF4 protein, which escapes degradation in the cytoplasm, is transported to the nucleus where it is functional. The results suggest that levels of MORF4 in cells must be tightly controlled and one mechanism involves stability of the protein.
authors.affiliation: Sam and Ann Barshop Institute for Longevity and Aging Studies UTHSCSA, STCBM, San Antonio, TX 78245, USA. tominaga@uthscsa.edu
authors.forename: Kaoru
authors.initials: K
authors.lastname: Tominaga
authors.forename: Emiko
authors.initials: E
authors.lastname: Tominaga
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Ausserlechner
authors.forename: Olivia M
authors.initials: OM
authors.lastname: Pereira-Smith
mesh_terms: D001565:Benzoates / Q000494:pharmacology; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D003513:Cycloheximide / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D004318:Doxycycline / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005663:Furans / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology; D010455:Peptides / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D061988:Proteasome Inhibitors; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D011720:Pyrazoles / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D025801:Ubiquitin / Q000235:genetics / Q000378:metabolism*; D044764:Ubiquitin-Activating Enzymes / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: C523934:4(4-(5-nitro-furan-2-ylmethylene)-3,5-dioxo-pyrazolidin-1-yl)-benzoic acid ethyl ester; D001565:Benzoates; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D005663:Furans; D007976:Leupeptins; C109882:MORF4 protein, human; C470833:MORF4L1 protein, human; D009842:Oligopeptides; D010455:Peptides; D061988:Proteasome Inhibitors; D011720:Pyrazoles; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D025801:Ubiquitin; D003513:Cycloheximide; C085706:FLAG peptide; D046988:Proteasome Endopeptidase Complex; D044764:Ubiquitin-Activating Enzymes; D004318:Doxycycline; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; C078846:epoxomicin