Examining the transcriptional response of overexpressing anthranilate synthase in the hairy roots of an important medicinal plant Catharanthus roseus by RNA-seq

BMC Plant Biology

Research Article

Jiayi Sun¹  Christie A. M. Peebles¹  Cheng Sun²  Harish Manmathan³ 

[1] Chemical and Biological Engineering Department, Colorado State University, Campus delivery 1370, 80523, Fort Collins, USA;Department of biology, Colorado State University, 1878 Campus Delivery, 80521, Fort Collins, Colorado, USA;Key Laboratory of Pollinating Insect Biology of the Ministry of Agriculture, Institute of Apicultural Research, Chinese Academy of Agriculture Science, 10093, Beijing, China;Soil and Crop Sciences Department, Colorado State University, Campus deliver 1170, 80523, Fort Collins, Colorado, USA;

关键词: Terpenoid indole alkaloid;    Madagascar periwinkle;    Transcription factors;    Plant secondary metabolism;    High-throughput sequencing;    Plant stress response;   

DOI  :  10.1186/s12870-016-0794-4

 received in 2015-10-29, accepted in 2016-04-29,  发布年份 2016

来源: Springer

【 摘 要 】

BackgroundClinically important anti-cancer drugs vinblastine and vincristine are solely synthesized by the terpenoid indole alkaloid (TIA) pathway in Catharanthus roseus. Anthranilate synthase (AS) is a rate-limiting enzyme in the TIA pathway. The transgenic C. roseus hairy root line overexpressing a feedback insensitive ASα subunit under the control of an inducible promoter and the ASβ subunit constitutively was previously created for the overproduction of TIAs. However, both increases and decreases in TIAs were detected after overexpressing ASα. Although genetic modification is targeted to one gene in the TIA pathway, it could trigger global transcriptional changes that can directly or indirectly affect TIA biosynthesis. In this study, Illumina sequencing and RT-qPCR were used to detect the transcriptional responses to overexpressing AS, which can increase understanding of the complex regulation of the TIA pathway and further inspire rational metabolic engineering for enhanced TIA production in C. roseus hairy roots.ResultsOverexpressing AS in C. roseus hairy roots altered the transcription of most known TIA pathway genes and regulators after 12, 24, and 48 h induction detected by RT-qPCR. Changes in the transcriptome of C. roseus hairy roots was further investigated 18 hours after ASα induction and compared to the control hairy roots using RNA-seq. A unigene set of 30,281 was obtained by de novo assembly of the sequencing reads. Comparison of the differentially expressed transcriptional profiles resulted in 2853 differentially expressed transcripts. Functional annotation of these transcripts revealed a complex and systematically transcriptome change in ASαβ hairy roots. Pathway analysis shows alterations in many pathways such as aromatic amino acid biosynthesis, jasmonic acid (JA) biosynthesis and other secondary metabolic pathways after perturbing AS. Moreover, many genes in overall stress response were differentially expressed after overexpressing ASα.ConclusionThe transcriptomic analysis illustrates overexpressing AS stimulates the overall stress response and affects the metabolic networks in C. roseus hairy roots. The up-regulation of endogenous JA biosynthesis pathway indicates the involvement of JA signal transduction to regulate TIA biosynthesis in ASαβ engineered roots and explained why many of the transcripts for TIA genes and regulators are seen to increase with AS overexpression.

【 授权许可】

CC BY   
© Sun et al. 2016