Identification of microRNAs from Medicinal Plant Murraya koenigii by High-Throughput Sequencing and Their Functional Implications in Secondary Metabolite Biosynthesis.

Claudia Gutiérrez-García,Shiek S S J Ahmed,Aashish Srivastava,Sathishkumar Ramalingam,Dhivya Selvaraj,Sujay Paul,Ashutosh Sharma

doi:10.3390/plants11010046

Abstract

MicroRNAs (miRNAs) are small noncoding RNA molecules that play crucial post-transcriptional regulatory roles in plants, including development and stress-response signaling. However, information about their involvement in secondary metabolism is still limited. Murraya koenigii is a popular medicinal plant, better known as curry leaves, that possesses pharmaceutically active secondary metabolites. The present study utilized high-throughput sequencing technology to investigate the miRNA profile of M. koenigii and their association with secondary metabolite biosynthesis. A total of 343,505 unique reads with lengths ranging from 16 to 40 nt were obtained from the sequencing data, among which 142 miRNAs were identified as conserved and 7 as novel miRNAs. Moreover, 6078 corresponding potential target genes of M. koenigii miRNAs were recognized in this study. Interestingly, several conserved and novel miRNAs of M. koenigii were found to target key enzymes of the terpenoid backbone and the flavonoid biosynthesis pathways. Furthermore, to validate the sequencing results, the relative expression of eight randomly selected miRNAs was determined by qPCR. To the best of our knowledge, this is the first report of the M. koenigii miRNA profile that may provide useful information for further elucidation of the involvement of miRNAs in secondary metabolism. These findings might be crucial in the future to generate artificial-miRNA-based, genetically engineered M. koenigii plants for the overproduction of medicinally highly valuable secondary metabolites.

Highlights

Murraya koenigii (L.) Spreng (Rutaceae) is a subtropical medicinal plant native to Asia and distributed throughout the subcontinent of India [1]
In this study, utilizing high-throughput Illumina sequencing technology, a total of 8,186,145 raw reads were obtained from the M. koenigii leaf tissue samples
The present study showed that both conserved and novel M. koenigii miRNAs were implicated in the terpenoid backbone biosynthesis, which can be synthesized either by the mevalonate pathway (MVA) in the cytoplasm or by the 2C-methyld-erythritol-4-phosphate pathway (MEP) in the plastids [47]

Summary

Introduction

Murraya koenigii (L.) Spreng (Rutaceae) is a subtropical medicinal plant native to Asia and distributed throughout the subcontinent of India [1]. Husna et al [7] observed that the ethanolic extract of curry leaves induced an antihyperglycemic effect in nicotinamide–streptozotocin-induced diabetic rats by decreasing the malondialdehyde level, associated with the generation of free radicals, and increasing the GSH level. These and other potential pharmacological activities of M. koenigii extracts are principally attributed to their secondary metabolites such as alkaloids, terpenoids, and flavonoids, especially their major constituents, the carbazole alkaloids [8]. It was noticed that mahanine caused cell cycle arrest in glioma cancerous cells by the downregulation of M-phase inducer phosphatase 3 (Cdc25c), cell division cycle protein 2 homolog (Cdc2), and cyclin B1 [10]

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Conclusion