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The maize dek1 gene functions in embryonic pattern formation and cell fate specification

Philip W. Becraft1,2,3,*, Kejian Li1,3, Nrisingha Dey1 and Yvonne Asuncion-Crabb1

1 Zoology and Genetics Department, Iowa State University, Ames, IA 50011, USA
2 Agronomy Department, Iowa State University, Ames, IA 50011, USA
3 Molecular Cellular and Developmental Biology Program, Iowa State University, Ames, IA 50011, USA



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  		Fig. 1. Phenotypes of dek1 mutant kernels. (A) dek1-388 mutant kernel. (B) dek1-D mutant kernel. (C) Medial cut through a wild-type (left) and dek1-1394 mutant (right) kernel. (D) Wild-type endosperm and aleurone. (E) dek1-792 mutant endosperm. (F,G) Section of a wild-type embryo and endosperm at (F)7 DAP and (G) 12 DAP. (H,I) Section of a dek1-792 embryo and endosperm at (H) 7 DAP and (I) at 12 DAP. Black arrows indicate embryos; white arrows designate transfer cells. Scale bars: 100 µm (D,E); 200 µm (F,H); 500 µm (G,I).

 


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  		Fig. 3. Generation of dek1 genetic mosaics and examples of an autonomous phenotype. (A) Diagram depicting the strategy for generating plants genetically mosaic for the dek1-792 allele. (B) Fluorescence micrograph of a leaf with mutant epidermal cells over wild-type internal cells. The normal mesophyll cannot rescue the mutant phenotype in the epidermis. The sector caused the leaf to bend, highlighting the influence of the epidermis on leaf morphology. (C) Another fluorescence micrograph of a sector. In the mesophyll, lack of red-fluorescing chloroplasts indicates that the tissue is genetically mutant for dek1. The guard cells are the only epidermal cells to contain chloroplasts. (D) Enlargement of the boxed region shown in C. The red-fluorescing wild-type guard cell (arrowhead) is directly adjacent to an epidermal cell showing a clear mutant phenotype (arrow). (E) Mutant mesophyll cells in a section of leaf with wild-type epidermis. The cells show unusual lobing. (F) Section where the internal mesophyll and bundle sheath cells are wild type (arrows) and the outer mesophyll layer is mutant and shows a mutant phenotype. The epidermis is wild type except for the bulged region between the arrowheads on the adaxial (upper) surface. (G) A single epidermal cell with mutant phenotype (asterisk) surrounded on 3 sides by normal looking cells. The phenotype appears cell-autonomous. (H) A cell with wild-type phenotype (asterisk) surrounded on 3 sides by cells with mutant phenotypes. The arrowhead designates the boundary between abnormal and normal bulliform cells. (I) An example of a cupped hair. Scale bars 100 µm (B-D,F,I); 50 µm (E); 10 µm (G,H).

 


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  		Fig. 2. The dek1-D mutant transforms leaf epidermal cells to bulliform cells. (A) dek1-D mutant plant. (B) Ear from a dek1-D mutant, showing a barren region in the middle of the inflorescence. (C,D) Cross sections of (C) wild-type leaf with bulliform cells (arrow) and (D) dek1-D mutant leaf. (E) SEM of a wild-type leaf showing a bulliform row (arrows). (F) SEM of dek1-D. (H) High magnification of bulliform cells on a wild-type leaf. (H) High magnification of dek1-D epidermal cells showing characteristics of bulliform cells. (I,J) High magnification of epidermal ground cells and a stoma in wild type (I) and dek1-D (J). Scale bars: 100 µm (C-F); 10 µm (G-J).

 


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  		Fig. 4. The mutant phenotype sometimes appears non cell-autonomous. (A) A pair of mutant sectors (brackets) with some cells between showing a mild mutant phenotype. The boxed region is shown in higher magnification in B. The arrows indicate cells partially affected by surface ridges typical of mutant cells. (C) Light and (D) fluorescence micrographs of a sector where a wild-type mesophyll cell is surrounded on 3 sides by mutant cells. The wildtype cell has a large spherical morphology not typical of normal mesophyll cells. (E) Wild-type mesophyll cells (between arrows) subjacent to a mutant epidermal sector showed a mutant phenotype. Size bars 100 µm (A,C,D); 50 µm (B,E).

 


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  		Fig. 5. Ectopic margin associated with a mutant sector. (A) The margin formed on the proximal side of the sector. (B) Fluorescence micrograph of the ectopic margin. The midrib is toward the right of the figure and the normal margin toward the left. (C,D), Scanning electron micrographs of the ectopic margin and the neighboring mutant cells. The boxed region in C is shown magnified in D. The arrow indicates epidermal cells with a mutant phenotype on the distal side of the ectopic margin. Scale bars 50 µm (B), 100 µm (C,D).

 


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  		Fig. 6. dek1-D, crinkly4-1231 double mutant analysis. (A,B) Scanning electron micrographs of a typical cr4-1231 single mutant. Arrows in A denote a bulliform row. The boxed area is enlarged in B. The arrows in B indicate cells with mild bulliform-like surface ridges. Note the aberrant cell shapes and the mismatched crenulations of neighboring cell sidewalls. (C) A dek1-D single mutant. Some cells appear normal, reflecting the leaky allele and mild expression. (D) A double mutant showing primarily the dek1-D phenotype. Most epidermal cells show the surface features of bulliform cells and lack the irregular morphology of cr4. (E) Over-expanded rectangular cells that lack interlocking crenulations along their sidewalls. (F) Highly irregular cells on a double mutant with a strong phenotype. (G) Cross section of a cr4-1231 single mutant. (H) A typical cross section through a double mutant. (I) A cross section through a strong double mutant. The arrow highlights an example of apparent multiple epidermal cell layers. (J) RT-dependent PCR amplification of Cr4 transcripts from wild-type and dek1-792 mutant endosperm. Lane 1, dek1-792 mutant endosperm, with reverse transcriptase (+RT); lane 2, wild-type endosperm (+RT); lane 3, Cr4 cDNA plasmid template; lane 4, dek1-792 RNA (—RT); lane 5, wild-type RNA (—RT). Scale bars 100 µm (A-D,G-I); 10 µm (E,F).

 

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