Os02g0743400
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Annotated Information
Function
OsPIN1 was expressed in the vascular tissues and root primordial in a manner similar to AtPIN1. Adventitious root emergence and development were significantly inhibited in the OsPIN1 RNA interference (RNAi) transgenic plants, which was similar to the phenotype of NPA (N-1-naphthylphalamic acid, an auxintransport inhibitor)-treated wild-type plants. α-naphthylacetic acid (α-NAA) treatment was able to rescue the mutated phenotypes occurring in the RNAi plants. Overexpression or suppression of the OsPIN1 expression through a transgenic approach resulted in changes of tiller numbers and shoot/root ratio. Taken together, these data suggest that OsPIN1 plays an important role in auxindependent adventitious root emergence and tillering[1][2][3].
Five adventitious roots were formed at the coleoptile node of 7-day-old seedlings in both the RNAi transgenic and the wild-type plants. In the second week, numbers of adventitious roots developed from the first node of the wildtype plants, while no new adventitious roots were initiated until the third week in the RNAi transgenic plants. Moreover, the number of adventitious roots in the RNAi transgenic plants was significantly less than in the wild-type plants.
Results showed that adventitious root primordia were initiated in 7-day-old seedlings of both wild-type and OsPIN1 RNAi transgenic plants, indicating that the formation of adventitious root primordia at the first node was not affected by the underexpression of OsPIN1. However, in the 14-day-old seedlings, new adventitious roots had developed in the wildtype plants, while they were not visible in the OsPIN1 RNAi transgenic plants. The adventitious root primordial in these RNAi plants stayed at the same size as those in the 7-day-old seedlings, indicating that the emergence of adventitious root primordia was arrested in these RNAi seedlings. Data showed that the development of adventitious roots in RNAi seedlings ceased at the sixth stage.
Expression
The expression of OsPIN1 in different tissues was analyzed by reverse transcriptase PCR (RT-PCR) and a β-glucuro- nidase (GUS) assay of transgenic plants expressing an OsPIN1 promoter/GUS fusion construct. RT-PCR analysis showed that OsPIN1 mRNA was transcribed in all tissues tested. The expression of the gene was higher in leaf, flower and seed than in root, stem and root collar. To confirm the expression patterns, a binary vector containing the GUS gene driven by the OsPIN1 promoter was constructed and used for rice transformation. The transgenic plants showed GUS expression in the vascular tissues of root, stem, anther, leaf and embryo of the seed. Cross-sections showed that GUS is also expressed in the primordia of adventitious and lateral roots, suggesting that OsPIN1 may be involved in root development. To investigate the function of OsPIN1 in rice, transgenic plants overexpressing or underexpressing OsPIN1 were produced using 35S-driven OsPIN1 and OsPIN1 RNA interfering (RNAi) constructs. Two transgenic lines were selected to represent the overexpressing or underexpressing groups. They were designated as 35S1 and 35S2, and RNAi1 and RNAi2, respectively. At the seedling stage, 14 days after germination, plant height, primary root length, adventitious root number and lateral root number on the primary root were investigated in wild-type and transgenic plants. The tiller number was counted at the maximum tillering stage (80 days after germination) under field conditions. The results showed that overexpression of the OsPIN1 gene significantly increased the primary root length and lateral root number, while underex- pression of OsPIN1 resulted in a reduced number of adventitious roots and a significantly increased number of tillers. Moreover, the tiller angles in the RNAi plants exceeded 30 degrees, which was much higher than the wildtype plants.
To verify whether OsPIN1 is involved in polar auxin transport, seedlings of the wild-type and transgenic plants were exposed to 0.1 μM α-naphthylacetic acid (α-NAA) and 0.5 μM NPA. The numbers of adventitious roots at the 7, 14 and 21- day-old seedling stages were recorded (Table 2). Results showed that the suppression of the OsPIN1 gene inhibited the formation of adventitious roots. Thus the number of adventitious roots in the OsPIN1 RNA interference (RNAi) transgenic plants was significantly less than that in wild-type plants (Table 2). NPA treatment had a similar effect on the wild-type plants (Table 2), suggesting that underexpression of the OsPIN1 gene may have affected the auxin transport process. Exogenously applied NAA on the RNAi transgenic plants partially complemented the inhibition effect of the underexpression of OsPIN1, confirming that OsPIN1 is involved in polar auxin transport.
Figure show that the expression level of OsPIN1 was inversely correlated with the sensitivity of the plants to the NPA treatment. In NPA treatment, the adventitious root numbers in transgenic plants overexpressing OsPIN1 (35S1and 35S2) were significantly higher than in wild-type seedlings. This suggests that the higher level of OsPIN1 expression had partially overcome the inhibition effect of the NPA treatment on the formation of adventitious roots. On the other hand, the OsPIN1 RNAi plants were hypersensitive to the treatment of NPA. Application of NPA to the OsPIN1 RNAi transgenic seedlings resulted in these seedlings losing their apical dominance in shoot growth, and eventually dying after a 14-day treatment. These results provided further evidence that OsPIN1 is involved in polar auxin transport.
Evolution
OsPIN1, previously named REH1, was proposed as an auxin transporter and a member of the PIN2 protein family. The full-length REH1 cDNA contains an open reading frame (ORF) of 1,785 nucleotides. The ORF encodes a protein of 595 amino acid residues with a calculated molecular mass of 65 kDa. Alignment between the cDNA and the genomic sequence of REH1 indicates that the REH1 gene has five introns. The deduced amino acid sequence of REH1 shows 69% sequence identity with Arabidopsis AtPIN1. Hydropathy and transmembrane motif analyses of the deduced amino acid sequence revealed that the REH1 protein contains three characteristic regions, including a hydrophobic region with five transmembrane segments, a predominantly hydrophilic core, followed by another hydrophobic region with five transmembrane segments. This is consistent with the results of Luschnig et al. (1998). Alignment of members of the PIN1 family across different species revealed that the highly conserved portions of the family are uniformly located in the transmembrane domains. Phylogenetic analysis of 32 members of PIN family showed that REH1 was evolutionarily closer to the PIN1 family than to the PIN2 family. It was renamed as OsPIN1.
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Labs working on this gene
State Key Laboratory of Plant Physiology and Biochemistry, College of Life Sciences, Zhejiang University, Kaixuan Road, Hangzhou, Zhejiang 310029, PR China
References
- ↑ Xu M, Zhu L, Shou H, Wu P. A PIN1 family gene, OsPIN1, involved in auxin-dependent adventitious root emergence and tillering in rice. Plant Cell Physiol. 2005 Oct;46(10):1674-81.
- ↑ Benková E, Michniewicz M, Sauer M, Teichmann T, Seifertová D, Jürgens G, Friml J. Local, efflux-dependent auxin gradients as a common module for plant organ formation. Cell. 2003 Nov 26;115(5):591-602.
- ↑ Blilou I1, Xu J, Wildwater M, Willemsen V, Paponov I, Friml J, Heidstra R, Aida M, Palme K, Scheres B.The PIN auxin efflux facilitator network controls growth and patterning in Arabidopsis roots. Nature. 2005 Jan 6;433(7021):39-44.
