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First published online 21 September 2005
doi: 10.1242/dev.02012

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AUXIN RESPONSE FACTOR1 and AUXIN RESPONSE FACTOR2 regulate senescence and floral organ abscission in Arabidopsis thaliana

¹ Department of Biology, University of North Carolina at Chapel Hill, CB #3280, Coker Hall, Chapel Hill, NC 27599-3280, USA
² Department of Biochemistry, University of Missouri, Columbia, Columbia, MO 65211, USA
³ Biology Department, Western Washington University, Bellingham, WA 98225-9160, USA

View larger version (15K): [in a new window]   Fig. 1. Identification of ARF1 and ARF2 T-DNA insertions. (A) Positions of T-DNA insertions in ARF1 and ARF2. Boxes represent exons, lines represent non-coding regions, and inverted triangles indicate T-DNA insertions. The region used for creation of a dsRNAi construct is indicated by a bar. Arrowheads indicate the locations of primers used to create probes. (B) Northern blot analysis of ARF gene expression in T-DNA insertion mutants and dsARF2 plants. RNA was extracted from 10-day-old seedlings.

View larger version (122K): [in a new window]   Fig. 2. Natural senescence in wild-type and arf2 rosette leaves. Leaves and inflorescences from 8-week-old plants are laid out in order of emergence.

View larger version (25K): [in a new window]   Fig. 3. Dark-induced senescence in detached wild-type and mutant leaves. (A) Chlorophyll levels in detached leaves following an 8 day incubation in the dark. Black bars represent chlorophyll levels just after detachment and white bars represent chlorophyll levels of leaves incubated in the dark for 8 days. Error bars indicate s.d. (n=6). (B) Time course of chlorophyll loss in Columbia and arf2-8 plants. Black circles indicate chlorophyll levels of Columbia leaves and white circles indicate arf2-8 leaves. (C) Northern blot analysis of gene expression in detached leaves. Leaves from Columbia and arf2-8 plants were detached, placed in darkness for various time periods, and then used for chlorophyll measurement or RNA extraction. Error bars indicate s.d. (n=6).

View larger version (158K): [in a new window]   Fig. 4. GUS::ARF2 protein levels during dark-induced senescence. PARF2::GUS::ARF2 plants (Li et al., 2004) were grown under short day conditions for 6 weeks. Mature, fully expanded leaves (A-D) or young, expanding leaves (E,F) were detached and stained immediately (A,E) or were placed in darkness for 2 days (B), 4 days (C,F) or 6 days (D) prior to staining.

View larger version (77K): [in a new window]   Fig. 5. Effects of arf gene mutations on flower development. Stage 13 Columbia (A) and arf2-8 flowers (E). Primary inflorescences of Columbia (B), Ws-0 (C) arf1-5 arf2-8 (F) and arf1-4 dsARF2 (G) plants. Abscission zones of Columbia (D) and arf1-5 arf2-8 (H) floral organs. St, stamen; P, petal; Se, sepal. Arrowheads indicate the abscission zone.

View larger version (62K): [in a new window]   Fig. 6. Analysis of ARF and IAA promoter activity in Arabidopsis flowers. PARF1::GUS (A-C) and PARF2::GUS (D-F) activity in flower bunches (A,D), stage 13 flowers (B, E) and stage 17 flowers (C, F). PIAA3::GUS (G-L) and PIAA7::GUS (M-R) activity in stage 13 flowers (G,I,K,M,O,P) and stage 17 flowers (H,J,L,N,P,R) in wild-type (G,H,M,N), arf1-4 (I,J,O,P) and arf1-4 arf2-8 (K,L,Q,R) plants. (S) Northern blot analysis of IAA gene mRNA in seedlings and flowers. Seedlings were treated for 2 hours with 1 µM IAA or with a mock control.

View larger version (35K): [in a new window]   Fig. 7. Effects of ethylene and cytokinin signaling on arf2-8 plants. (A) Chlorophyll levels in detached leaves incubated in the dark for 8 days. Black bars represent chlorophyll levels just after detachment and white bars represent chlorophyll levels of leaves incubated in the dark for 8 days. Error bars indicate s.d. (n=6). (B) Average position at abscission where position 1 equals the first opened (stage 13) flower and flowers are numbered basipetally. Error bars indicate s.d. (n>20). (C) Northern blot analysis of ethylene-induced gene expression of arf2-8 plants. Six-week-old plants were incubated in airtight containers with either 10 ppm ethylene or air. RNA was extracted from whole rosettes. (D) Chlorophyll levels in cytokinin-treated detached leaves. Detached leaves were floated on water supplemented with 100 nM 6-benzyladenine or on a DMSO mock control. White, Columbia; cross hatched, Columbia+BA; black, arf2-8; hatched, arf2-8+BA. Error bars indicate s.d. (n=6).

View larger version (18K): [in a new window]   Fig. 8. Phenotypes of arf2-8 nph4-1 arf19-4 plants. (A) Chlorophyll levels in detached leaves incubated in the dark for 8 days. Black bars represent chlorophyll levels just after detachment and white bars represent chlorophyll levels of leaves incubated in the dark for 8 days. Error bars indicate s.d. (n=6). (B) Average position at abscission where position 1 equals the first opened (stage 13) flower and flowers are numbered basipetally. Error bars indicate s.d. (n=25). (C) Northern blot analysis of NPH4/ARF7 and ARF19 expression in senescing leaves. Leaves from Columbia and arf2-8 plants were detached, placed in the dark for various time periods and used for mRNA extraction.