Unveiling novel targets of paclitaxel resistance by single molecule long-read RNA sequencing in breast cancer

Bi Lian,Zhi-Ming Shao,Xin Hu

doi:10.1038/s41598-019-42184-z

Abstract

RNA sequencing has become one of the most common technology to study transcriptomes in cancer, whereas its length limits its application on alternative splicing (AS) events and novel isoforms. Firstly, we applied single molecule long-read RNA sequencing (Iso-seq) and de novo assembly with short-read RNA sequencing (RNA-seq) in both wild type (231-WT) and paclitaxel resistant type (231-PTX) of human breast cancer cell MDA-MBA-231. The two sequencing technology provide both the accurate transcript sequences and the deep transcript coverage. Then we combined shor-read and long-read RNA-seq to analyze alternative events and novel isoforms. Last but not the least, we selected BAK1 as our candidate target to verify our analysis. Our results implied that improved characterization of cancer genomic function may require the application of the single molecule long-read RNA sequencing to get the deeper and more precise view to transcriptional level. Our results imply that improved characterization of cancer genomic function may require the application of the single molecule long-read RNA sequencing to get the deeper and more precise view to transcriptional level.

Highlights

With the advancing technology of RNA sequencing (RNA-seq)[1], the mRNA level of variation can be clearly elaborated by known genes and their different transcriptions, which the number of transcription is 4–5 times than the genes[2]
Iso-seq is widely used in constructing genome reference of different species, which further more expending its application into high-throughput sequencing with de novo genome reference when combining with RNA-seq.[8]
Numerous researches have performed sequencing on genetic level to analyze genome profile of drug resistance

Summary

Introduction

With the advancing technology of RNA sequencing (RNA-seq)[1], the mRNA level of variation can be clearly elaborated by known genes and their different transcriptions, which the number of transcription is 4–5 times than the genes[2]. With the development of cancer processing, various number of unannotated AS events happened and formed novel transcription, which may play a critical role in cancer metastasis and drug resistance[6]. Iso-seq is widely used in constructing genome reference of different species, which further more expending its application into high-throughput sequencing with de novo genome reference when combining with RNA-seq.[8]. Increasing researches have remarkable founding on the function of some alternative splicing event and transcripts played a pivotal role the tumorigenesis, progression, and metastasis of cancer, as well as inducing drug resistance[14,15,16]. We treated one of the TNBC cell lines, MDA-MB-231 with paclitaxel to generate paclitaxel-resistant cells We used both RNA sequencing and Pacbio sequencing technology to perform paclitaxel resistant cells and www.nature.com/scientificreports/. We verified a novel isoform of BAK1 genes

Methods

Results

Conclusion