Dataset Information

Human HeLa cell lince SILAC - The arginylation branch of the N-end rule pathway positively regulates cellular autophagic flux and clearance of proteotoxic proteins

ABSTRACT: The arginylation branch of the N-end rule pathway positively regulates cellular autophagic flux and clearance of proteotoxic protein

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell

DISEASE(S): Malignant Neoplasm Of Ovary

SUBMITTER: Joon won Lee

LAB HEAD: Min Jae Lee

PROVIDER: PXD003813 | Pride | 2016-09-27

REPOSITORIES: Pride

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Publications

The arginylation branch of the N-end rule pathway positively regulates cellular autophagic flux and clearance of proteotoxic proteins.

Jiang Yanxialei Y Lee Jeeyoung J Lee Jung Hoon JH Lee Joon Won JW Kim Ji Hyeon JH Choi Won Hoon WH Yoo Young Dong YD Cha-Molstad Hyunjoo H Kim Bo Yeon BY Kwon Yong Tae YT Noh Sue Ah SA Kim Kwang Pyo KP Lee Min Jae MJ

Autophagy 20160825 11

The N-terminal amino acid of a protein is an essential determinant of ubiquitination and subsequent proteasomal degradation in the N-end rule pathway. Using para-chloroamphetamine (PCA), a specific inhibitor of the arginylation branch of the pathway (Arg/N-end rule pathway), we identified that blocking the Arg/N-end rule pathway significantly impaired the fusion of autophagosomes with lysosomes. Under ER stress, ATE1-encoded Arg-tRNA-protein transferases carry out the N-terminal arginylation of ...[more]

PMID: 27560450

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Project description:Activation of the serine/threonine kinase Akt contributes to the formation, maintenance, and therapeutic resistance of cancer, which is driving development of compounds that inhibit Akt. Phosphatidylinositol ether lipid analogues (PIAs) are analogues of the products of PI3K that inhibit Akt activation, translocation, and the proliferation of a broad spectrum of cancer cell types. To gain insight into the mechanism of PIAs, time-dependent transcriptional profiling of 5 active PIAs and the PI3K inhibitor LY294002 (LY) was performed in non-small cell lung cancer (NSCLC) cells using high-density oligonucleotide arrays. Gene ontology analysis revealed genes involved in apoptosis, wounding response, and angiogenesis were upregulated by PIAs, while genes involved in DNA replication, repair and mitosis were suppressed. Genes that exhibited early differential expression were partitioned into 3 groups; those induced by PIAs only (DUSP1, KLF6, CENTD2, BHLHB2, PREX1), those commonly induced by PIAs and LY (TRIB1, KLF2, RHOB and CDKN1A), and those commonly suppressed by PIAs and LY (IGFBP3, PCNA, PRIM1, MCM3 and HSPA1B). Increased expression of the tumor suppressors RHOB (RhoB), KLF6 (COPEB) and CDKN1A (p21Cip1/Waf1) was validated as an Akt-independent effect that contributed to PIA-induced cytotoxicity. Despite some overlap with LY, active PIAs have a distinct expression signature that contributes to their enhanced cytotoxicity. H157 cells were plated at 2x10^6 in T-75 flasks in RPMI medium 1640 containing 10% FBS and incubated for 24h. The media was then changed to RPMI medium 1640 with 0.1% FBS and the cells were incubated overnight. The following morning, cells were treated with 10 microM PIA6 dissolved in DMSO for 0h, 2h, 6h or 12h, and an equal volume of DMSO was added to control samples. For the PIA comparison, 10 microM PIAs (5, 6, 7, 23, 24, 25) or 10 microM LY294002 (LY) were incubated with the cells for 6h. In the PIA comparison experiment, the same concentration of PIA7 (an inert analog consisting of only the lipid side chain) was used as a control. Following incubation, the alterations in cellular morphology were photographed, and cells from 6-well plates were harvested for immunoblot analysis. Total RNA was extracted from cells treated in T-75 flasks using TRIzol reagent (Invitrogen) and chloroform, and purified according to the RNeasy midiprep spin kit protocol (Qiagen). Oligonucleotide microarray was performed with dye-swap. Microarray chips were generated from the 34,580 longmer probe set Human Genome Oligo Set Version 3.0 (Qiagen). Protocols for cDNA labeling, hybridization, and scanning are available through the National Human Genome Research Institute microarray core.

Dataset Information

Human HeLa cell lince SILAC - The arginylation branch of the N-end rule pathway positively regulates cellular autophagic flux and clearance of proteotoxic proteins

Publications

The arginylation branch of the N-end rule pathway positively regulates cellular autophagic flux and clearance of proteotoxic proteins.

Similar Datasets

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