The nuclear genome of Rhazya stricta and the evolution of alkaloid diversity in a medically relevant clade of Apocynaceae.

Jamal S M Sabir,Emmanuel Noutahi,Virginie Calderon,Dhivya Arasappan,Tracey A Ruhlman,Nahid H Hajrah,Abdullah Y Obaid,Chunfang Zheng,Abdulrahman L Al-Malki,David Sankoff,Robert K Jansen,Mohammed N Baeshen,Nabih A Baeshen,Meshaal J Sabir,Seongjun Park,Nadia El-Mabrouk,Mohammad A Khiyami

doi:10.1038/srep33782

Abstract

Alkaloid accumulation in plants is activated in response to stress, is limited in distribution and specific alkaloid repertoires are variable across taxa. Rauvolfioideae (Apocynaceae, Gentianales) represents a major center of structural expansion in the monoterpenoid indole alkaloids (MIAs) yielding thousands of unique molecules including highly valuable chemotherapeutics. The paucity of genome-level data for Apocynaceae precludes a deeper understanding of MIA pathway evolution hindering the elucidation of remaining pathway enzymes and the improvement of MIA availability in planta or in vitro. We sequenced the nuclear genome of Rhazya stricta (Apocynaceae, Rauvolfioideae) and present this high quality assembly in comparison with that of coffee (Rubiaceae, Coffea canephora, Gentianales) and others to investigate the evolution of genome-scale features. The annotated Rhazya genome was used to develop the community resource, RhaCyc, a metabolic pathway database. Gene family trees were constructed to identify homologs of MIA pathway genes and to examine their evolutionary history. We found that, unlike Coffea, the Rhazya lineage has experienced many structural rearrangements. Gene tree analyses suggest recent, lineage-specific expansion and diversification among homologs encoding MIA pathway genes in Gentianales and provide candidate sequences with the potential to close gaps in characterized pathways and support prospecting for new MIA production avenues.

Highlights

Alkaloid production in plants is limited in distribution and the specific alkaloid repertoire is variable across taxa
Using genomic and transcriptomic sequences from Rauvolfioideae along with species in the order Gentianales and beyond, we identified homologs of characterized monoterpenoid indole alkaloid (MIA) pathway genes revealing the lineage specific nature of their evolution
Comparison to assembly statistics reported for three recently published nuclear genomes demonstrates the high quality of the Rhazya draft (Supplementary Table S4)

Summary

Results and Discussion

We sequenced three Illumina libraries yielding nearly 400 million reads to generate approximately 112X coverage of the R. stricta ~274 Mb genome. The two Rhazya sequences and RG were 76% and 77% identical whereas identities to Rauvolfia SGD were 58% and 57% at the amino acid level It appears that the R. stricta genome may not encode SGD, our own mass spectrometry analysis (Supplementary Table S9) along with decades of reports demonstrate the production of MIAs that derive from the reactive strictosidine aglycone, a product of SGD activity. Much like the other MIA pathway genes examined, THAS family members showed a pattern of lineage specific expansion in the Rauvolfioideae and included two sequences from R. stricta in the reduced tree (Fig. 3, Supplementary Fig. S2). The gene phylogeny analyses facilitated by the Rhazya sequences combined with available data indicate lineage specific expansion resulting in distinct sets of MIA enzyme homologs in the Rauvolfioideae as well as in the Gentianales. A critical evaluation of this question would require more than a single representative from each lineage to draw meaningful conclusions

Materials and Methods

Author Contributions

Additional Information