Post-Embryonic Nerve-Associated Precursors to Adult Pigment Cells: Genetic Requirements and Dynamics of Morphogenesis and Differentiation
Shown are analyses of time-lapse movies for trunks derived from wild-type and mutant larvae (N = 281 larvae, 5241 total cells examined). (A) Total numbers of newly arising hypodermal mitfa::GFP+ cells differed among genotypes (square root transformed data, F5,273 = 30.2, P<0.0001). Shown are least squares means (±95% confidence intervals) after controlling for significant differences among stages (F2,273 = 3.9, P<0.0001) and normalized to cells mm−2 day−1. Letters above bars indicate means that differed significantly (P<0.05) by Tukey-Kramer post hoc comparisons. Numbers within bars indicate numbers of larval trunks examined. (B) The origins of new hypodermal mitfa::GFP+ cells differed among genotypes (χ2 = 145.6, d.f. = 10, P<0.0001; n = 1582 total new cells). Bar widths are proportional to the total numbers of new hypodermal cells observed in each genotype (shown in A). DV, cells entering the hypodermis after migrating over the dorsal or ventral myotome margins; HM, cells entering from the vicinity of the horizontal myoseptum. M, cells entering from within the myotomes. The sources of mitfa::GFP+ cells did not differ significantly across stages overall (χ2 = 0.003, d.f. = 4, P = 1), though different genotypes exhibited stage-dependent variation (stage x genotype interaction: χ2 = 46.3, d.f. = 20, P<0.0001; not shown). (C) Frequencies of differentiation, death and proliferation differed among genotypes. Bar widths are proportional to the total numbers of hypodermal mitfa::GFP+ cells and melanophores observed per larva, after controlling for area and duration of imaging, and normalized to cells mm−2 day−1 (larva means±SE: wild-type, 116±9; erbb3b, 13±8; tuba8l3a, 62±9; kita, 72±10; csf1r, 78±14; ednrb1, 61±16). Differentiation, The likelihood of mitfa::GFP+ cells acquiring melanin during imaging differed among genotypes (χ2 = 100.6, d.f. = 5, P<0.0001; n = 335 total differentiating cells): the relatively few erbb3b and tuba8l3a mutant cells were especially likely to differentiate whereas very few kita mutant cells differentiated. Additional effects were attributable to stage (χ2 = 30.3, d.f. = 2, P<0.0001) and a stage x genotype interaction (χ2 = 27.3, d.f. = 10, P<0.0001; not shown). Death, The incidence of mitfa::GFP+ cells dying during imaging differed among genotypes (χ2 = 116.9, d.f. = 5, P<0.0001; n = 507 total dying cells) with particularly high rates of death in kita and ednrb1 mutants. Additional variation was attributable to differences among stages (χ2 = 23.5, d.f. = 2, P<0.0001) and a stage x genotype interaction (χ2 = 20.4, d.f. = 10, P<0.05) resulting from an increased likelihood of ednrb1 mutant cells dying at later stages (not shown) Division, The incidence of mitfa::GFP+ cells dividing differed significantly among genotypes (χ2 = 23.6, d.f. = 5, P<0.0001; n = 142 total dividing cells). Asymmetric confidence intervals, Bayesian 95% upper and lower bounds.