IKU1Gene Encodes A VQ Motif Protein Controlling Seed Size In Arabidopsis

Seed represents an important source of food, feed, and industrial raw material for mankind. Seed size is one of the significant factors affecting crop production. Endosperm accounts for most of the bulk of cereal seed and determines the seed size. It will be strategically significant for the sustainable development of the agricultural production and the human society to support the research on reproductive biology, because of the resource shortage and expanding population in the world.The reproductive process in plants is an important area in biology, which still requires substantial genetic and molecular investigation to elucidate the complex process. Angiosperms, or higher plants, have a unique seed forming process, double fertilization: the egg cell is fused with a sperm nucleus and the central cell fuses to another sperm nucleus both coming from the same pollen grain. This leads to the formation of the embryo and endosperm. Three mutants ikul, iku2and mini3have been shown to give small seed size because of the reduction of endosperm growth. The expression levels of MINI3and IKU2were decreased in the ikul-1mutant. IKU2expression was reduced in a miniS-1background whereas MINI3expression levels were unaltered in the iku2-3mutant. These observation suggest the successive action of the three genes IKU1, IKU2and MINI3in the same genetic pathway of seed development. In this pathway, MINI3encodes WRKY10, a WRKY class transcription factor; IKU2encodes a LRR KINASE (At3g19700). But the identity of IKU1remains unknown.In this study, the integrated approaches involving classic genetic, molecular, cellular methodologies were used to investigate the molecular identity of IKU1. We reported here the cloning of IKU1gene which controls the seed size in Arabidopsis using map-based strategy. It was shown that IKU1is At2g35230which encodes a novel protein containing VQ motif. VQ motif contains a core sequence of unknown function and is specific in plant species. The genetic complementation of ikul phenotype suggests that IKU1gene functions in endosperm. It was also confirmed that the endosperm and the embryo of Arabidopsis thaliana are independently transformed through infiltration by Agrobacterium tumefaciens. The mutation in IKU1only causes obvious phenotype in seed, though IKU1is expressed widely. The genomic construct carrying mutations in VQ motif failed to complement the ikul seed lesion, suggesting an essential role of the VQ motif in IKU1functioning. There is evidence that the mutations in other parts of genes are not important for IKU1function. We found that IKU1is nuclear-localized, organized into speckles and led to gene silencing under35S. When driven by its native promoter, the GFP::IKU1without any speckle, able to complement the mutant phenotype, was detected in the nuclear of the early endosperm as well as other vegetative organs. The physical interaction between IKU1and MINI3/WRKY10suggests that the IKU1is a co-transcription factor of MTNI3/WRKY10. A model has been proposed to explain how the IKU1and MINI3/WRKY10regulate endosperm development and seed size. In wild type Arabidopsis, IKU1interacts with MINI3/WRKY10to form a complex which regulates the IKU2and consequently wild-type seeds will be produced. Seeds in ikul or miniS mutants become small, because IKU2is repressed. Further, in ikul mutants the IKU1-MINI3/WRKY10transcription complex becomes unstable and the un-associated MINI3/WRKY10protein starts to repress the expression of MINIS genes as a feedback mechanism. In the same time IKU2expression is down-regulated, which leads the formation of small seeds.