Figure 1.
A. Leaf rosettes of five-week-old wild type and abs5-1D mutant. To have a clear view of the rosette leaves, the inflorescence stems were removed prior to photographing. B–C. Overview of the transverse sections of the eighth rosette leaf from three-week-old wild type (B) and abs5-1D (C). Bars: 500 µm. D–E. Transverse sections of the mid-vein regions of wild type (D) and abs5-1D (E) leaf. Bars: 50 µm.
Table 1.
Comparison of the average number of secondary inflorescences of wild type, abs5-1D and ABS5/T5L1 OE lines.
Figure 2.
A. Co-segregation analysis of abs5-1D. Genomic DNAs were extracted from 16 F2 progenies from a cross between wild type and abs5-1D. DNA samples were digested with HindIII and separated on an agarose gel before transferred onto a nylon membrane. The DNA gel blot was probed with 32P-labelled BAR gene sequences. Plants with abs5-1D-like phenotypes were marked with asterisks. B. Schematic representation of the T-DNA insertion site in abs5-1D. Open boxes represent genes in the vicinity of the activation tagging T-DNA. Solid lines represent intergenic regions. The orientations of these genes' open reading frames (ORFs) were indicated by arrows. C. Accumulation of At1g68810 transcripts in wild-type and abs5-1D. Equal amounts of total RNA (5 µg) extracted from 2-week-old seedlings were separated on a formaldehyde gel and transferred to a nylon membrane. The blot was hybridized with 32P-labelled full-length At1g68810 cDNA. The ethidium bromide-stained RNA gel served as a loading control. D. Leaf rosettes of representative five-week-old wild type and two independent At1g68810 over-expression lines (OE-1 and OE-2). E. Semi-quantitative RT-PCR analysis of At1g68810 transcripts accumulation in the plants shown in (C).
Figure 3.
Expression analysis of ABS5/T5L1.
A. Semi-quantitative RT-PCR analysis of ABS5/T5L1 gene expression in various tissues: roots, two-week-old seedlings, rosette leaves, stems, cauline leaves, siliques and flowers. The expression of Actin2 was used as a control. B. Nuclear localization of ABS5-GFP fusion protein in Arabidopsis leaf protoplasts. Wild type leaf protoplasts were transformed with P35S::GFP or P35S::ABS5-GFP. A single representative protoplast was shown for each transformation. Nuclei were specifically stained by the fluorescent dye Hoechst33342. Hoechst33342, GFP, chlorophyll autofluorescence signals were monitored by fluorescence microscopy. Bright field (BF) images served as controls for protoplast integrity. Bars: 10 µm.
Figure 4.
Comparisons of DR5::GUS activities and cotyledon vein patterns between wild type and abs5-1D.
A–D. DR5::GUS activities in heterozygous DR5::GUS (A and B) and DR5::GUS abs5-1D/+ double heterozygous backgrounds (C and D). Illustrated are representative cotyledons (A and C) and the first true leaves (B and D) from two-week-old seedlings. E–G. DR5::GUS activities in the first true leaves of two-week-old homozygous DR5::GUS lines (E) and two independent 35S promoter driven ABS5/T5L1 OE lines in DR5::GUS background (F and G). H. Representative cotyledon vein patterns in wild type. Cotyledons with two, three, or four areoles were observed in wild type [35]. I. Quantification of cotyledon vein patterns in 10-day-old wild type (n = 423) and abs5-1D (n = 412). J. Abnormal cotyledon vein patterns observed in abs5-1D. Illustrated are cotyledons with abnormal tertiary veins (top row) and cotyledons with five areoles (bottom row).
Table 2.
Quantification of cotyledon vein patterns in wild type and abs5-1D.
Figure 5.
A. Phenotypes of representative two-week-old wild type and abs7-1D seedlings. B. Comparison of individual leaves detached from plants shown in (A). From left to right are the two cotyledons and the first four rosette leaves (Upper panel: wild type; Lower panel: abs7-1D). C. Comparison of the overall plant statues of three-week-old wild type, abs5-1D and abs7-1D. D-F. Transverse sections of the ninth rosette leaves of three-week-old wild type (D), abs5-1D (E) and abs7-1D (F). G–I. Transverse sections of the first rosette leaves of three-week-old wild type (G), abs5-1D (H) and abs7-1D (I).
Figure 6.
A. Co-segregation analysis of abs7-1D. 16 randomly selected plants from the F2 progeny from a cross between wild type and abs7-1D were used for analysis. Southern blot analysis was performed as in Figure 2A. Plants with abs7-1D-like phenotypes were marked by asterisks. B. Schematic representation of the T-DNA insertion site in abs7-1D. Genes in the vicinity of the T-DNA insertion site were represented by white boxes. Arrows indicated the orientation of the ORFs of these genes. C. Phenotypes of representative three-week-old wild type and two ABS7 over-expression lines (OE-8 and OE-10). D. Expression levels of ABS7 gene in wild type and two ABS7 OE lines analyzed via semi-quantitative RT-PCR. E. Semi-quantitative RT-PCR analysis of ABS7 gene expression in roots, two-week-old seedlings, rosette leaves, stems, cauline leaves, siliques and flowers. The expression of Actin2 was used as a control. F. Nuclear localization of ABS7-GFP fusion protein in Arabidopsis leaf protoplasts. Protoplast transformation, nucleus staining and fluorescence microscopy was performed as in Figure 3B.
Figure 7.
Transcriptional activation analysis of ABS5/T5L1 and ABS7/MYB101 in yeast.
Yeast strain AH109 was transformed with a negative control vector (pBD), a positive control pBD-WRKY33, pBD-ABS5 or pBD-ABS7, respectively. Each of the BD vectors was co-transformed with an empty AD vector, pGADT7. A–C. Growth of yeast transformants on the SD/-Trp-Leu medium (A), the SD/-Trp-Leu-His medium plus 5 mM 3-AT (B) or the SD/-Trp-Leu-His-Ade medium (C). D. Activation of the LacZ gene analyzed via filter lifting X-β-gal assays.
Figure 8.
Effects of epidermal specific expression of ABS5/T5L1 or ABS7/MYB101.
A. Verification of the epidermal specific AtML1 promoter (PAtML1). The transverse sections of young leaves from the PAtML1::GFP expressing Arabidopsis lines were examined via con-focal microscopy. Mesophyll cells were visualized through the chlorophyll autofluorescence. B. Phenotypes of three-week-old wild type and two independent PAtML1::ABS5 lines. C. Phenotypes of two-week-old wild type and two independent PAtML1::ABS7 lines. D–F. DR5::GUS activities in the first true leaves of two-week-old homozygous DR5::GUS line (D) and two independent lines expressing PAtML1::ABS5 in DR5::GUS background (E and F).