Transcriptomic findings light on mechanisms of marine and terrestrial evolutionary adaptation of Rhizophoraceae plants

Rhizophoraceae has 2 terrestrial genera and 4 marine genera. The intertidal zone in which marine mangroves are located is known for its low oxygen and high salinity. Marine and terrestrial genera have evolved distinct adaptive characteristics, among which viviparous reproduction is the most unique. To investigate the genetic foundations difference underlying the adaptive mechanisms of marine–terrestrial genera, we selected two species from Rhizophoraceae. Kandelia obovata is marine and viviparous, and Carallia brachiata is terrestrial and non-viviparous. We compared their transcriptome of 8 tissues (root, stem, leaf, flower, ovule, fruit, seed, embryo) and found that the mature reproductive organs (fruit, seed, embryo) of K. obovata did not reduce metabolic activity compared to C. brachiata. The reproductive organs of K. obovata were regulated by the same gene set as vegetative organs. This contrasted with C. brachiata. Eight kinds of hormone transduction genes were up-regulated in the seed of K. obovata. Finally, and most importantly, the transcriptional factors AP2 and ARF families were significantly more expressed in the reproductive organs of K. obovata than in those of C. brachiata. At the same time, the ERF family was more expressed in its roots. The findings suggested that the hormone transduction may contribute to viviparous initiation. Transcriptional factors were quite crucial for mangroves' adaptation to wetlands.

Methods
Reverse transcription (RT) was performed with 1 μg total RNA isolated from three independent biological replicates per species using 5X All-In-One RT MasterMix (ABM, Canada). The reaction procedure is as follows: incubate at 25 °C for 10 minutes, then incubate at 42 °C for another 15 minutes, and inactivate the reaction at 85 °C for 5 minutes. First-strand cDNA was diluted to 200 ng/μL as a quantitative template. The relative quantification was done by ChamQTM Universal SYBR® qPCR Master Mix (Vazyme, China). The reaction system was 10 μL 2 x SYBR Green PCR Mix, 0.5 μL upstream primer, 0.5 μL downstream primer, 8 μL H2O. RT-qPCR was performed on the above mixture, and the reaction procedure was as follows: 95 °C, 30 s; 95 °C, 10 s; 58 °C, 10 s; 72 °C, 20 s, and 40 cycles. Actin was selected for the reference gene. The relative expression of the target gene was calculated by the 2−ΔΔCt method.