Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

CMTM6 drives cisplatin resistance by regulating AKT mediated Wnt signaling

ABSTRACT: The RNA-seq library preparation was performed for H357 CisR Nt Sh and H357 CisR CMTM6 shRNA treated with vehicle control (UT) and cisplatin (10 μM) (CisT) for 24 h. We have included two independent biological replicates to identify the CMTM6 downregulation mediated global transcriptome changes. For RNA-seq library preparation 2 μg of total RNA was used to isolate mRNA through magnetic beads using mRNA isolation kit (PolyA mRNA isolation Module, NEB) followed by RNA-seq library preparation using mRNA library preparation kit (NEB) strictly following the vendor recommended protocol. After library preparation concentration of libraries was estimated using qubit 2.0 (Invitrogen) and the recommended fragmentation sizes were confirmed by Tapestation (Agilent).

INSTRUMENT(S): NextSeq 550

ORGANISM(S): Homo sapiens

SUBMITTER: Rupesh Dash

PROVIDER: E-MTAB-9424 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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CMTM6 drives cisplatin resistance by regulating Wnt signaling through the ENO-1/AKT/GSK3β axis.

Mohapatra Pallavi P Shriwas Omprakash O Mohanty Sibasish S Ghosh Arup A Smita Shuchi S Kaushik Sandeep Rai SR Arya Rakesh R Rath Rachna R Das Majumdar Saroj Kumar SK Muduly Dillip Kumar DK Raghav Sunil K SK Nanda Ranjan K RK Dash Rupesh R

JCI insight 20210222 4

Rewiring tumor cells to undergo drug-induced apoptosis is a promising way to overcome chemoresistance. Therefore, identifying causative factors for chemoresistance is of high importance. Unbiased global proteome profiling of sensitive, early, and late cisplatin-resistant oral squamous cell carcinoma (OSCC) lines identified CMTM6 as a top-ranked upregulated protein. Analyses of OSCC patient tumor samples demonstrated significantly higher CMTM6 expression in chemotherapy (CT) nonresponders as comp ...[more]

PMID: 33434185

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Project description:Objectives: The sequencing by the PolyA selection is the most common approach for library preparation. With limited amount or degraded RNA, alternative protocols such as the NuGEN have been developed. However, it is not yet clear how the different library preparations affect the downstream analyses of the broad applications of RNA sequencing. Methods and Materials: Eight human mammary epithelial cell (HMEC) lines with high quality RNA were sequenced by Illumina’s mRNA-Seq PolyA selection and NuGEN ENCORE library preparation. The following analyses and comparisons were conducted: 1) the numbers of genes captured by each protocol; 2) the impact of protocols on differentially expressed gene detection between biological replicates; 3) expressed single nucleotide variant (SNV) detection; 4) non-coding RNAs, particularly lincRNA detection; and 5) intragenic gene expression. Results: Sequences from the NuGEN protocol had lower (75%) alignment rate than the PolyA (over 90%). The NuGEN protocol detected fewer genes (12-20% less) with a significant portion of reads mapped to non-coding regions. A large number of genes were differentially detected between the two protocols. About 17-20% of the differentially expressed genes between biological replicates were commonly detected between the two protocols. Significantly higher numbers of SNVs (5-6 times) were detected in the NuGEN samples, which were largely from intragenic and intergenic regions. The NuGEN captured fewer exons (25% less) and had higher base level coverage variance. While 6.3% of reads were mapped to intragenic regions in the PolyA samples, the percentages were much higher (20-25%) for the NuGEN samples. The NuGEN protocol did not detect more known non-coding RNAs such as lincRNAs, but targeted small and “novel” lincRNAs. Conclusion: Different library preparations can have significant impacts on downstream analysis and interpretation of RNA-seq data. The NuGEN provides an alternative for limited or degraded RNA but it has limitations for some RNA-seq applications. Eight human mammary epithelial cell (HMEC) lines with high quality RNA were sequenced by Illumina’s mRNA-Seq PolyA selection and NuGEN ENCORE library preparation. The goal was to look the impacts of protocols on results and intepretation.

Dataset Information

CMTM6 drives cisplatin resistance by regulating AKT mediated Wnt signaling

Publications

CMTM6 drives cisplatin resistance by regulating Wnt signaling through the ENO-1/AKT/GSK3β axis.

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