The Molecular Mechanism Of ARF10 And ARF16 Regulated Root Development In Arabidopsis

Root indeterminate growth and its outstanding ability to produce new tissues continuously make this organ a highly dynamic structure able to respond promptly to external enviromental stimuli.It is reported that environmental cues can greatly influence root development through the regulation of auxin synthesis,transport and signaling transduction and finally protect themselves from various environmental stress.Previous studies showed that auxin response factor 10(ARF10)and 16(ARF16),the components of auxin signaling pathway,play important roles in the maintenance of stem cell niche in root tip and regulation of lateral root development.However,the molecular mechanism of ARF10 and ARF16 regulating root development in Arabidopsis remain elusive.In the present study,we make efforts to determine the role of ARF10 and ARF16 in regulating stem cell niche in Arabidopsis.The following conclusions can be drawn from our experiments:ARF10 and ARF16 overexpression plants showed that starch granules accumulate in all columella layers including cells at the position of the columella stem cells,indicating that ARF10 and ARF16 overexpression plants fails to maintain root stem cell.In contrast,arf10/16 double mutant has tumor-like stem cell phenotype.These results indicated that ARF10 and ARF16,especially ARF10,play important roles in stem cell niche maintance in root.Next,we analyze the expression profiling of arfl0/16 double mutant,and we found three genes(IAAn,STOP2 and TAT3)were significantly induced or suppressed in the double mutant,suggesting that these three genes may be involved in the process of root growth and development in Arabidopsis regulated by ARF10 and ARF16.Our qRT-PCR also showed that IAAn gene was induced in arf10 and arf16,especially in arf10/16 double mutant account for that ARF10 and ARF16 inhibit the expression of IAAn.The subsequent ChIP experiment show that ARF10 and ARF16 protein is in combination with promoter of IAAn gene at-2102 and-1093 auxin response elements,and inhibit the expression of IAAn.The yeast two hybrid and BiFC experiment verify that IAAn could interact with ARF10 and ARF16,and the interaction of IAAn and ARF10 is strong.It suggested that IAAn combine with ARF10 may regulate the expression of IAAn itself by forming a feedback inhibition.Our qRT-PCR also showed that STOP2 gene was suppressed in arf10 and arfl6,especially in arf10/16 double mutant account for that ARF10 and ARF16 promote the expression of STOP2.Two mutant strains of STOP2 showed two layers of stem cell phenotype,consisting with the phenotype of arf10/16 double mutant,indicating that ARF10 and ARF16 induce cell differentiation by activating the expression of STOP2.These results provide new clues for further study on root development regulation by auxin signaling pathway involving ARF10 and ARF16 in Arabidopsis.