vvi-miPEP172b and vvi-miPEP3635b increase cold tolerance of grapevine by regulating the corresponding MIRNA genes.

As a kind of small molecular weight proteins, many peptides have been discovered, including peptides encoded by pri-miRNA (miPEPs). Similar as traditional phytohormone or signaling molecular, these peptides participate in numerous plant growth processes. MicroRNAs (miRNAs) play an important regulatory role in plant stress response. While the roles of miPEPs in response to abiotic stress has not been studied now. In this study, to explore whether miPEPs could contribute to low temperature (4ºC) tolerance of plants, the expression pattern of 23 different vvi-MIRs were analyzed by qRT-PCR in 'Thompson Seedless' (Vitis vinifera) plantlets under cold stress (4ºC) firstly, and vvi-MIR172b and vvi-MIR3635b which showed an elevated expression levels were selected to identify miPEPs. Through transient expression, one small open reading frame (sORF) in each of the two pri-miRNAs could increase the expression of corresponding vvi-MIR, and the amino acid sequences of sORFs were named vvi-miPEP172b and vvi-miPEP3635b, respectively. The synthetic vvi-miPEP172b and vvi-miPEP3635b were applied to the grape plantlets, and the tissue culture plantlets exhibited a higher cold tolerance compared with the control groups. These results revealed the effective roles of miPEPs in plant cold stress resistance for the first time, providing a theoretical basis for the future application of miPEPs to agricultural production.

A miRNA-Encoded Small Peptide, vvi-miPEP171d1, Regulates Adventitious Root Formation.

One of the biggest challenges in clonal propagation of grapevine (Vitis vinifera) is difficulty of rooting. Adventitious root initiation and development are the critical steps in the cutting and layering process of grapevine, but the molecular mechanism of these processes remains unclear. Previous reports have found that microRNA (miRNA)-encoded peptides (miPEPs) can regulate plant root development by increasing the transcription of their corresponding primary miRNA. Here, we report the role of a miPEP in increasing adventitious root formation in grapevine. In this study, we performed a global analysis of miPEPs in grapevine and characterized the function of vvi-miPEP171d1, a functional, small peptide encoded by primary-miR171d. There were three small open reading frames in the 500-bp upstream sequence of pre-miR171d. One of them encoded a small peptide, vvi-miPEP171d1, which could increase the transcription of vvi-MIR171d Exogenous application of vvi-miPEP171d1 to grape tissue culture plantlets promoted adventitious root development by activating the expression of vvi-MIR171d Interestingly, neither exogenous application of the vvi-miPEP171d1 peptide nor overexpression of the vvi-miPEP171d1 coding sequence resulted in phenotypic changes in Arabidopsis (Arabidopsis thaliana). Similarly, application of synthetic ath-miPEP171c, the small peptide encoded by the Arabidopsis ortholog of vvi-MIR171d, inhibited the growth of primary roots and induced the early initiation of lateral and adventitious roots in Arabidopsis, while it had no effect on grape root development. Our findings reveal that miPEP171d1 regulates root development by promoting vvi-MIR171d expression in a species-specific manner, further enriching the theoretical research into miPEPs.