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Fig. 7. Quantitative analyses of melanophore morphogenesis in D. rerio and
D. albolineatus. (A) In regions where melanophore stripes develop in
D. rerio, melanophore numbers are dramatically reduced in D.
albolineatus and this deficit becomes more pronounced through pigment
pattern metamorphosis. (B) In the middle of the flank, where the first
interstripe region forms in D. rerio, melanophore numbers are
increased in D. albolineatus, reflecting the absence of a distinctive
and persistent melanophore-free region in the anterior of the flank. Despite
the increased melanophore numbers, the overall complement of melanophores is
dramatically reduced overall compared with D. rerio. (C) Analyses of
twice daily image series reveal differential appearance ('births') and
disappearance ('deaths') of melanophores between species. Shown are cumulative
mean births and deaths recorded in each of three developing larvae through
middle stages of pigment pattern metamorphosis, with `births' defined as the
appearance of new melanophores (either by differentiation or proliferation)
and `deaths' defined by the unambiguous loss of melanophores (see Materials
and methods). Melanophore births were not significantly different between
species (F1,107=1.87, P=0.2) after controlling
for variation among individuals (F4,107=2.72,
P<0.05) and across days (F21,107=5.43,
P<0.001). By contrast, melanophore deaths were significantly
greater in D. albolineatus than D. rerio
(F1,81=3.04, P<0.005) after controlling for
variation among individuals (F4,81=2.10, P=0.09)
and across days (F29,81=2.47, P<0.001). Error
bars are omitted for clarity. (D) Total melanophore movements are reduced in
D. albolineatus compared with D. rerio, both during early
and middle metamorphosis (left), and through later metamorphosis (right).
Shown are mean (±s.e.m.) distances moved by individual melanophores,
with distances expressed as percentages of the flank height. Left, species
differences were significant (F1,696=14.23,
P<0.0005; n=368, 344 melanophores in D. rerio
and D. albolineatus) after controlling for variation associated with
individuals (nested within species: F4,696=2.60,
P<0.05), and anteroposterior region nested within individuals
(F10,696=5.19, P<0.0001). Right, species
differences were significant (F1,708=8.45,
P<0.005; n=291, 419 melanophores in D. rerio and
D. albolineatus) whereas inter-individual differences were not
significant (P=0.2). (E,F) Directional movements of melanophores were
significantly reduced in D. albolineatus compared with D.
rerio. Each point represents a single melanophore followed from its first
appearance to its final position at the end of the series or until it was lost
(n=368, 344 melanophores in D. rerio and D.
albolineatus, respectively). Plots show the initial dorsoventral
positions at which melanophores first appeared, and the subsequent changes in
dorsoventral positions by the end of the images series. The dorsal-most
position on the flank is assigned a relative value of 0, and the ventral-most
position on the flank is assigned a value of 1. Regression slopes are
estimated separately for dorsal and ventral regions of the flank because of
differences in shape and growth. (E) In D. rerio, melanophores that
initially appear in more dorsal regions of the flank tend to move ventrally
whereas melanophores that initially appear in more ventral regions of the
flank tend to move dorsally (partial regression coefficients±s.e.m. for
relationship between starting dorsoventral position and arcsine-transformed
movements for dorsal and ventral regions of the flank, respectively:
-0.17±0.02, -0.14±0.04). (F) In D. albolineatus,
directional movements were significantly reduced compared with D.
rerio in dorsal regions of the flank (F1,399=6.43,
P<0.05), although species differences were not detectable in
ventral regions (F1,399=1.40, P=0.2), as assessed
by the magnitude of starting position xspecies interactions [partial
regression coefficients for dorsal and ventral, respectively:
-0.10±0.02, -0.10±0.02; after controlling in both dorsal and
ventral analyses for variation among individuals (nested within species, both
P<0.005), variation among the three examined anteroposterior
regions of the flank nested within individuals (both P<0.0005),
main effects of species (P=0.7, P<0.005, respectively),
and starting position independent of species (both P<0.0001)].
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