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Proteomics of HeLa cells starved and treated with YWF or leptomycin B

ABSTRACT: HeLa cells labeled with SILAC and were amino acids starved and treated with YWF, leptomycin B, MG132 or chloroquine. Cells were lysed with a 8M Urea buffer and proteins were reduced and alkylated. Equal amount of proteins of different labels were mixed and were digested with trypsin and resulting peptides were injected on a Q Exactive plus instrument. Raw data were analysed with MaxQuant with default settings.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Bertrand Fabre

PROVIDER: MSV000092564 | MassIVE | Tue Aug 01 06:27:00 BST 2023

REPOSITORIES: MassIVE

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Publications

Regulation of nucleo-cytosolic 26S proteasome translocation by aromatic amino acids via mTOR is essential for cell survival under stress.

Livneh Ido I Cohen-Kaplan Victoria V Fabre Bertrand B Abramovitch Ifat I Lulu Chen C Nataraj Nishanth Belugali NB Lazar Ikrame I Ziv Tamar T Yarden Yosef Y Zohar Yaniv Y Gottlieb Eyal E Ciechanover Aaron A

Molecular cell 20230901 18

The proteasome is responsible for removal of ubiquitinated proteins. Although several aspects of its regulation (e.g., assembly, composition, and post-translational modifications) have been unraveled, studying its adaptive compartmentalization in response to stress is just starting to emerge. We found that following amino acid starvation, the proteasome is translocated from its large nuclear pool to the cytoplasm-a response regulated by newly identified mTOR-agonistic amino acids-Tyr, Trp, and P ...[more]

PMID: 37738964

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Gene Expression in Cisplatin Resistance HeLa Cells

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Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis Methods: RNA from 14-day-old seedlings of wild-type, brip1, brip2, brm-3, brm-1, brip1 brip2, brip1 brip2 brm-3 and brip1 brip2 brm-1 was isolated using the RNAprep Pure Plant Kit as described above. RNAs from three biological replicates was sequenced separately at Novogene, using Illumina Hiseq X-Ten (Sequencing method: Hiseq-PE150). Results: Reads were mapped to the TAIR10 Arabidopsis genome using TopHat (Galaxy V2.1.1 in usegalaxy.org) with default settings, except that a minimum intron length of 20 bp and a maximum intron length of 4,000 bp were required (Paired-end). Then, mapped reads were assembled according to TAIR10 version of genome annotation using cufflinks (version 2.1.1) with default settings. To analyze differential expression, the assembled transcripts from three independent biological replicates in Col and other mutants were included and compared using Cuffdiff (version 2.1.1) with default settings. Conclusions: Our study represents the first detailed analysis of brip1, brip2, brip1 brip2 transcriptomes, with biologic replicates, generated by RNA-seq technology.

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Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis Methods: RNA from 14-day-old seedlings of wild-type, brip1, brip2, brm-3, brm-1, brip1 brip2, brip1 brip2 brm-3 and brip1 brip2 brm-1 was isolated using the RNAprep Pure Plant Kit as described above. RNAs from three biological replicates was sequenced separately at Novogene, using Illumina Hiseq X-Ten (Sequencing method: Hiseq-PE150). Results: Reads were mapped to the TAIR10 Arabidopsis genome using TopHat (Galaxy V2.1.1 in usegalaxy.org) with default settings, except that a minimum intron length of 20 bp and a maximum intron length of 4,000 bp were required (Paired-end). Then, mapped reads were assembled according to TAIR10 version of genome annotation using cufflinks (version 2.1.1) with default settings. To analyze differential expression, the assembled transcripts from three independent biological replicates in Col and other mutants were included and compared using Cuffdiff (version 2.1.1) with default settings. Conclusions: Our study represents the first detailed analysis of brip1, brip2, brip1 brip2 transcriptomes, with biologic replicates, generated by RNA-seq technology.

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