title: Two Degradation Pathways of the p35 Cdk5 (Cyclin-dependent Kinase) Activation Subunit, Dependent and Independent of Ubiquitination.
abstract: Cdk5 is a versatile protein kinase that is involved in various neuronal activities, such as the migration of newborn neurons, neurite outgrowth, synaptic regulation, and neurodegenerative diseases. Cdk5 requires the p35 regulatory subunit for activation. Because Cdk5 is more abundantly expressed in neurons compared with p35, the p35 protein levels determine the kinase activity of Cdk5. p35 is a protein with a short half-life that is degraded by proteasomes. Although ubiquitination of p35 has been previously reported, the degradation mechanism of p35 is not yet known. Here, we intended to identify the ubiquitination site(s) in p35. Because p35 is myristoylated at the N-terminal glycine, the possible ubiquitination sites are the lysine residues in p35. We mutated all 23 Lys residues to Arg (p35 23R), but p35 23R was still rapidly degraded by proteasomes at a rate similar to wild-type p35. The degradation of p35 23R in primary neurons and the Cdk5 activation ability of p35 23R suggested the occurrence of ubiquitin-independent degradation of p35 in physiological conditions. We found that p35 has the amino acid sequence similar to the ubiquitin-independent degron in the NKX3.1 homeodomain transcription factor. An Ala mutation at Pro-247 in the degron-like sequence made p35 stable. These results suggest that p35 can be degraded by two degradation pathways: ubiquitin-dependent and ubiquitin-independent. The rapid degradation of p35 by two different methods would be a mechanism to suppress the production of p25, which overactivates Cdk5 to induce neuronal cell death.
authors.affiliation: From the Laboratory of Molecular Neuroscience and.
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Takasugi
authors.affiliation: From the Laboratory of Molecular Neuroscience and.
authors.forename: Seiji
authors.initials: S
authors.lastname: Minegishi
authors.affiliation: From the Laboratory of Molecular Neuroscience and.
authors.forename: Akiko
authors.initials: A
authors.lastname: Asada
authors.affiliation: From the Laboratory of Molecular Neuroscience and.
authors.forename: Taro
authors.initials: T
authors.lastname: Saito
authors.affiliation: Laboratory of Cellular Biochemistry, Department of Biological Sciences, and Graduate School of Sciences, Tokyo Metropolitan University, Mianami-osawa, Hachioji,Tokyo 192-0397, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kawahara
authors.affiliation: From the Laboratory of Molecular Neuroscience and hisanaga-shinichi@tmu.ac.jp.
authors.forename: Shin-ichi
authors.identifier: http://orcid.org/0000-0002-7572-472X
authors.initials: S
authors.lastname: Hisanaga
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D051360:Cyclin-Dependent Kinase 5 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004622:Embryo, Mammalian / Q000166:cytology; D004789:Enzyme Activation; D057809:HEK293 Cells; D006207:Half-Life; D006801:Humans; D054878:Lipoylation; D008813:Mice, Inbred ICR; D009154:Mutation; D009474:Neurons / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D010770:Phosphotransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D055550:Protein Stability; D059748:Proteolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D054875:Ubiquitination
chemical_list: C492236:Cdk5r1 protein, mouse; D011994:Recombinant Proteins; D010770:Phosphotransferases; D051360:Cyclin-Dependent Kinase 5; C495906:Cdk5 protein, mouse; D046988:Proteasome Endopeptidase Complex
keywords: cyclin-dependent kinase (CDK); cyclin-dependent kinase 5; degron; neuron; proteasome; protein degradation; serine/threonine protein kinase; ubiquitin