MiR156-mediated Regulatory Mechanism Of Chlorophyll Degradation In Rice

Chlorophyll,an important plant photosynthetic pigment,is the basis of photosynthesis.When green plant organs enter the senecent stage,chlorophyll content and photosynthetic capacity gradually decrease.Chlorophyll degradation is closely related to crop yield and quality.This process is regulated by not only the external cues,but also by endogenous coding genes.Therefore,the study of molecular mechanism of chlorophyll degradation in plants is critical for agricultural production.miR156 is an evolutionarily conserved miRNA in plants,and it plays an important role in regulating plant productivity by repressing its target genes,Squomosa Promoter Binding Protein-like genes.Previous studies showed that miR156 affects chlorophyll content in tobacco and Arabidopsis.Therefore,we hypothesized that miR156 might affect crop yields in part by regulating the chlorophyll degradation pathway To test this hypothesis,we generated mimicry156 transgenic rice plants with reduced level of miR156,and analyzed the effects of miR156 and SPLs on chlorophyll content and other yield-related traits.We uncovered a link between miR156 and chlorophyll degradation in rice.The results are as follows:1.A miR156 knocking-down construct was generated using a target mimicry strategy.it was transferred into rice by Agrobacterium-mediated genetic transformation method.In this study,a total of 82 transgenic rice plants with reduced level of miR156 were recovered.2.Phenotypic characterization of mimicry156 transgenic plants showed that the numer of tillers was reduced,whereas chlorophyll content and fertility were increased in mimicry156;mimicry156 plants also have more fully-filled bigger seeds than wild type.3.Analyses of chlorophyll content,photosynthetic parameters and chlorophyll fluorescence suggested that the stay-green phenotype of mimicry156 plants belongs to the B-type function.4.In vitro leaf experiment showed that the rate of chlorophyll degradation in mimicry156 plant was much slower than that in wild type.q RT-PCR indicated that the NYC1 gene,involved in chlorophyll degradation,was downregulated in mimicry156 plants.Bioinformatic analysis suggested that there are some SPL protein binding sites in NYC1 promoter.EMSA demonstrated that SPL14 protein,one target of miR156,could directly bind to the promoter of NYC1 to regulate its expression.Therefore,miR156 regulates chlorophyll degradation by repressing SPL14,thus downreulating NYC1 to slow down the rate of chlorophyll degradation in mimicry156 plants.In this study,we uncovered the missing link between developmental timing and chlorophyll degradation in rice,and it is of potential use for crop improvement.