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First published online 4 October 2006
doi: 10.1242/dev.02599

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Chromatin assembly factor CAF-1 is required for cellular differentiation during plant development

¹ Institute of Plant Sciences and Basel-Zurich Plant Science Center, ETH Zurich, 8092 Zurich, Switzerland.
² Department of Plant and Microbial Biology, University of California, Berkeley, CA 94720, USA.

View larger version (45K): [in a new window]   Fig. 1. Development of CAF-1 mutant seedlings. (A) Representative 9-day-old light-grown seedlings of CAF-1 mutants and their corresponding wild types. (B) After induction of germination by a 10-hour exposure to white light, seeds were kept in the dark at 23°C. Hypocotyl lengths of at least 20 seedlings were determined after 4 days and expressed relative to their corresponding wild types. Means±s.e.m. of at least three independent experiments are shown.

View larger version (17K): [in a new window]   Fig. 2. Hypocotyl cell size, number and hypocotyl diameter in CAF-1 mutant seedlings. Seedlings of CAF-1 mutants and their corresponding wild types were grown as described for Fig. 1B, except that the seedlings were grown for 9 days. (A) The size of epidermal cells of the hypocotyl was measured for 10 seedlings of the mutants and wild types (about 300 cells per genotype). (B) The total number of epidermal cells along the hypocotyl axis was estimated from the hypocotyl length and the average cell size. (C) The diameters of hypocotyls of CAF-1 mutants and wild-type seedlings were determined. For A and C, the means±s.e.m. of the relative differences between mutant and wild type from three to five independent experiments are shown.

View larger version (24K): [in a new window]   Fig. 3. Ploidy in CAF-1 mutant seedlings. Seedlings were grown as described for Fig. 2 or exposed to LD conditions for 9 days. (A) Ploidy was measured for seedlings grown under LD photoperiods. (B) Ploidy was measured for etiolated CAF-1 seedlings. For three to four independent preparations, areas of 2C (white), 4C (light gray), 8C (dark gray) and 16C peaks (black bars) were averaged. For all measurements, the s.d. was smaller than 4%.

View larger version (67K): [in a new window]   Fig. 4. Leaf development in CAF-1 mutants. (A) Jigsaw-like shape of epidermal cells in CAF-1 mutants. Scale bar: 100 µm. (B,D) Size of epidermal cells on the first and second (B), or third and fourth (D) rosette leaves of CAF-1 mutants. Shown are the means±s.e.m. for one representative experiment (n63). (C,E) Number of epidermal cells on rosette leaves. (C) Epidermal cell number of the first and second rosette leaf. Shown is the average (means±s.e.m.) of three (msi1-as: two) independent experiments. (E) Epidermal cell number of the third and fourth leaf. Shown are the means±s.e.m. of one experiment (n7). Asterisks indicate P-values (t-test) <0.05; double asterisks indicate P-values <0.005.

View larger version (21K): [in a new window]   Fig. 5. Trichome differentiation in CAF-1 mutants. (A,B) Trichomes from leaves of wild-type (Col) plants usually have three branches (A), whereas trichomes on fas1-4 leaves develop up to six branches (B). (C-F) Trichomes of wild-type (Col) and fas1-4 plants are single cells with one nucleus. (C,D) DAPI-stained trichomes of wild type (Col) and fas1-4, respectively. (E,F) Close-up view of nuclei in DAPI-stained trichomes of wild type (Col) and fas1-4, respectively. (G) Trichome branching for the first two rosette leaves of at least six plants per experiment. Shown are the average values of several independent experiments. Segments represent fractions of trichomes with 1 (white hatched), 2 (white), 3 (light gray), 4 (dark gray), 5 (black) and 6 (dark gray, hatched) branches. (H) Ploidy of trichome nuclei from the first two rosette leaves. DNA in trichome nuclei was stained with DAPI and the fluorescence intensity values were normalized to the fluorescence of guard cell nuclei from the same specimen; these are displayed relative to wild-type intensities. Shown are the average values of several independent experiments (means±s.e.m.). Double asterisks indicate P-values (t-test) <0.005.

View larger version (12K): [in a new window]   Fig. 6. Epistasis analysis of CAF-1 function for trichome branching. (A) Trichome branching in fas1-4, fas2-4, msi1-as and double mutants. Branching was quantified for all trichomes on the first two rosette leaves of at least six plants per experiment. Segments represent fractions of trichomes with 2 (white), 3 (light gray), 4 (dark gray), 5 (black) and 6 (dark gray, hatched) branches. (B) Trichome branching in fas2-1, gl3-1 and double mutants. Branching was quantified for all trichomes on the first two rosette leaves of at least six plants per experiment. Segments represent fractions of trichomes with 1 (white), 2 (light gray), 3 (dark gray), 4 (black) and 5 (dark gray, hatched) branches.