Genomics Of High-altitude Adaptation And Evolution Of Przewalskia Tangutica(Solanaceae)

Przewalskia tangutica is a unique endangered Tibetan medicine plant in China,belongs to tribe Hyoscyameae in Solanaceae,which mainly grows in the high mountain areas above 4,000 meters in Qinghai-Tibet Plateau(QTP).After fertilization,the calyx of the plant gradually elongates to form an innovative structure of "Chinese Lantern" that wraps the whole fruit,making it a typical representative of the alpine specialized adaptation phenomenon.Such species are called as the airball plants,but the special structure has only been reported in several distantly related genera of the Solanaceae.It remains unknown that how P.tangutica adapts to the harsh environment at high altitude,and what genetic changes underlie evolutionary origins of such an innovative "Chinese Lantern" fruits between unrelated genera.In this study,the first high-quality chromosomal-level reference genome was assembled and annotated for P.tangutica.In combination with the genome-resequencing 30 individuals,the high-altitude adaptation,demographic dynamics and convergent evolutions of innovative fruit traits were further discussed.The main findings are summarized as follows:(1)Based on Illumina,Nanopore and Hi-C sequencing data,a high-quality reference genome of P.tangutica was assembled,and the contig N50 was 17.50 Mb.After that,1610 contigs were successfully anchored onto 23 chromosomes.Based on ab initio prediction and homologous prediction,50,828 protein-coding genes were annotated.The repetitive sequence is about 2.52 Gb,accounting for 83.27% of the genome size,and the long terminal repeats(LTRs)has undergone recent expansion,which leads to the larger genome.(2)Comparative genomics analyses revealed that the P.tangutica has undergone three whole-genome duplication events,and an additional WGD event occurred in～9.69 Mya alone.The occurrence of this duplication event coincided with the extensive climate change in the QTP,retaining genes mainly responding to cold,UV radiation,and DNA damage repair,which successfully helped P.tangutica adapt to the arid alpine environment.Compared with the low-altitude species of the Solanaceae,multiple transcription factor families showed a significant expansion in P.tangutica,and most were found to be related to the abiotic stress response.All these expanded gene families may together enhance the stress responding ability.(3)The gametophytic self-incompatibility(GSI)syntenic locus to the other obligate outcrossing Solanaceae species was broken by the inserted LTR in P.tangutica.Changes were also found for the flower-specific expression of these homologous genes,and the linked GSI genes in this species.Such changes may have led to its selfcompatibility.Three deeply diverged lineages were found in the QTP distribution of this species,and gene flow between them was weak but continuous.Three lineages diverged and decreased their population sizes since the largest glaciations occurred in the QTP approximately 720–500 thousand years ago.In addition,one obvious hybrid population between two lineages were identified,suggesting that genetic exchanges between and within lineages still occur.Therefore,this facultative breeding system may benefit this species in terms of both reproductive assurance from self-pollination and gene flow from outcrossing to avoid inbreeding depression.(4)The likely ‘convergent’ origin of the “Chinese lantern” as the inflated calyx syndrome(ICS),an innovative trait was further explored.The cis-regulatory elements of the MADS box gene MPF2 in the upstream 1000 bp promoter region,through which MPF2 is negatively regulated by MPF3 in non-ICS species,have been lost in this species.Such losses release the high heterotopic expression of MPF2 in calyx,which further contribute to ICS in P.tangutica while this gene was repressed by MPF3 in other non-ICS species.This is very similar to that found for the “Chinese lantern’ genus Physalis in which all elements disappeared within the regulatory region.However,the sequence alignment of this promoter region across two ICS species showed no same and ‘convergent’ nucleotide mutations and phylogenetic signals.In addition,ICS is critical for alpine survival of P.tangutica through preventing UV-B radiation and ensuring seed development and promoting self-pollination.The results of ecological experiments and transcriptome analysis showed that the structure of "Chinese Lantern" could effectively prevent UV-B radiation damage and abiotic stress,ensure seed development and promote self-pollination.Therefore,the inflated calyx of P.tangutica is an effective adaptive strategy in alpine environments and might have been selected for abiotic environmental conditions as well as for enhancing pollination success.In conclusion,this study not only enriches the genomics research of the Solanaceae,and reveals the high-altitude adaptation and population history of one alpine plant in the family,but also highlight the importance of the non-similar non-coding sequence variations of cis-regulatory elements in leading to ‘convergent’ origin of the similar innovative phenotypic traits.