In this case, the total length of the cell cycle would not be affected. Conversely, if dMyc acts directly to prolong S and/or G2, then overexpressed Stg should shorten G2, but the cells should compensate by extending G1. If the extended S and G2 phases in dMyc-overexpressing cells reflect compensation for an accelerated G1, we reasoned that coexpression of Stg with dMyc should prevent the G2 elongation and simultaneously shorten the cell cycle. The mass doubling time was calculated as described in the Experimental Procedures, from the median clone area. The total number of pixels within each clone (the clone area) was measured and converted to μM 2, where 1 pixel = 0.782 μM 2. Right panels, analysis of mass doubling times (mass DT) of the cell clones represented on the left (Act>GFP and Act>dMyc clones are a subset of the clones on left). Although the distribution of clonal cell number is similar for control, Stg alone, and dMyc alone, the median number of cells expressing dMyc+Stg is larger, as a result of a faster cell DT. Cell doubling times (cell DT) were calculated from the median size of the clone. Clones are displayed as the distribution of clonal cell number, with the range reflecting the inherent variability of cell division rates within the wing disc. Left panels, cells in each clone were counted, and cell doubling times for each genotype are indicated. (B) Act>dMyc, Act>Stg, or Act>dMyc+Stg cell clones were induced at 72 hr and analyzed at 120 hr AED. Since wing size is strictly proportional to body size ( The reduced size of dmyc mutant wing disc cells and the adult wing cells indicates that dmyc is required for normal cell size. ![]() ![]() These data demonstrate that the cell size defects in dmyc wings are dmyc specific. Expression of a UAS-dMyc transgene under control of a Gal4 driver that is expressed ubiquitously in the wing disc completely rescued the cell size reduction in dmyc P0 wings and also rescued the length (99%) and width (96%) of their wings ( Table 1). Calculation of an approximate area (length × width) of the mutant wings suggests that dmyc P1 wings may also have fewer cells. Thus, the small dmyc P0 wing size is due to smaller cells. Furthermore, measurement of the length and width of mutant and wild-type wings showed that wings from dmyc P0 and dmyc P1 males are between 8% and 15% smaller in each dimension than controls ( Table 1). The cell area in both mutants was approximately 14% smaller than wild-type cells ( Table 1). We scored the number of trichomes within a precisely defined area of the wing blade from wild-type and dmyc adult males and found more cells per unit area in dmyc P0 and dmyc P1 mutant wings than in control wings. Trichome density is indicative of cell size in the wing. Each epidermal cell of a wing secretes a single hair, called a trichome. To determine whether the dmyc mutant cells remained small throughout development, we examined cell size in adult wings. (n), number of animals scored nd, not determined. Complementation tests of dmyc P0 with X chromosome deficiencies were carried out by crossing dmyc P0 animals to wild-type (WT) flies or to flies carrying the deficiencies noted, and dymc P0/Df progeny were scored for viability. Rescue of specific dmyc phenotypes by overexpression of dMyc cDNA was carried out by three means (see footnotes a–c). A total of 20 dmyc P0, 20 dmyc P1, 26 dmyc P0 C765>dMyc, and 15 control male wings were analyzed. For wing length, vein 3 was measured from its origin in the hinge to the distal wing tip. Wing width is the distance from the posterior margin (at vein 5) perpendicular to the anterior margin. There were 43 squares from dmyc P0 male wings, 31 each from dymc P1 and control male wings, and 34 from dymc P0 C765>dMyc male wings. The number of trichomes from a specific region next to the posterior crossvein was counted within a 0.05 mm square and converted to area/cell (μM 2). Wings from dmyc mutant adults were examined for trichome density to determine cell size (area). Viability is expressed as a percentage of the expected number.
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