Developmental and Cellular Basis of Vertical Bar Color Patterns in the East African Cichlid Fish Haplochromis latifasciatus

Abstract

The East African adaptive radiations of cichlid fishes are renowned for their diversity in coloration. Yet, the developmental basis of pigment pattern formation remains largely unknown. One of the most common melanic patterns in cichlid fishes are vertical bar patterns. Here we describe the ontogeny of this conspicuous pattern in the Lake Kyoga species Haplochromis latifasciatus. Beginning with the larval stages we tracked the formation of this stereotypic color pattern and discovered that its macroscopic appearance is largely explained by an increase in melanophore density and accumulation of melanin during the first 3 weeks post-fertilization. The embryonal analysis is complemented with cytological quantifications of pigment cells in adult scales and the dermis beneath the scales. In adults, melanic bars are characterized by a two to threefold higher density of melanophores than in the intervening yellow interbars. We found no strong support for differences in other pigment cell types such as xanthophores. Quantitative PCRs for twelve known pigmentation genes showed that expression of melanin synthesis genes tyr and tyrp1a is increased five to sixfold in melanic bars, while xanthophore and iridophore marker genes are not differentially expressed. In summary, we provide novel insights on how vertical bars, one of the most widespread vertebrate color patterns, are formed through dynamic control of melanophore density, melanin synthesis and melanosome dispersal.

Neural innervation as a potential trigger of morphological color change and sexual dimorphism in cichlid fish

“…To test whether and how the morphological color change in M. auratus can be explained by changes in chromatophore number, distribution and characteristics, we compared chromatophores in yellow and dark morph using light microscopy of whole-mount scale preparations. In line with previous descriptions for cichlids 7,38 , three types of chromatophores could be detected in both morphs: melanophores with black to dark brown pigmentation, xanthophores with yellow to orange pigmentation, and iridophores that produce iridescent/reflective colors ( Supplementary Fig. S1).…”

“…Photographs of M. auratus scales were captured on Leica DM6B upright microscope with a Leica DMC 2900 camera. We captured images in three different modes: brightfield for melanophore coverage and dispersed diameter, polarized light for iridophore coverage 7 , fluorescent with GFP filter for xanthophore coverage and dispersed diameter 80,81 . To count the number of melanophores and xanthophores, we treated the scales immediately after taking the above photos with 10 mg/ml adrenaline (SIGMA-ALDRICH) for 5 min at room temperature on microscope slides to aggregate the melanosomes which permit robust cell number quantification ( Supplementary Fig.…”

“…In vertebrates the color of the integument is shaped by the multilayered arrangement and interaction of cells with different pigmentary and structural properties 5 . Variations in the density, shape and properties of chromatophores and the intracellular organization of pigments and reflective molecules shapes the macroscopic appearance of the integument 6 , 7 . Several types of pigment-bearing and light-reflecting chromatophores have been identified in teleosts 8 .…”

Functional conservation and divergence of color‐pattern‐related agouti family genes in teleost fishes

“…RNA extraction, complementary DNA (cDNA) synthesis and RT‐qPCR were performed as previous described (Kratochwil et al, 2018; Liang et al, 2020). Briefly, dissected skin tissues were stored in RNAlater (Invitrogen).…”

Spotting genome-wide pigmentation variation in a brown trout admixture context

“…chromatophores; mainly melanophores, iridophores, leucophores and xanthophores) are distributed in the hypo- and the epidermis in fish, and mutational or other analyses allowed for increased understanding in the complex mechanisms of pigment cell fates during development and their resulting distribution (Kelsh et al, 2004, 2009; Kimura et al, 2014; Eom, Bain, Patterson, Grout, & Parichy, 2015; Nüsslein-Völlard & Singh, 2017; Parichy & Spiewak, 2015; Singh & Nüsslein-Völlard, 2015; Salis et al, 2018; Volkening 2020). Studies now extend to other fish species (Maan & Sfec, 2013; Irion & Nüsslein-Völlard, 2019) and to a large array of eco-evolutionary questions regarding the involvement of genes or regulatory pathways in fish pigmentation, its epistatic and pleiotropic nature, its modularity and its control, sexually antagonist selection or its role in speciation as well as the impact of whole-genome duplication that promoted the diversification of pigment cell lineages (Hultman, Bahary, Zon, & Johnson, 2007; Miller et al, 2007; Braasch, Brunet, Volff, & Schartl, 2009; Roberts, Ser & Kocher, 2009; Albertson et al, 2014; Santos et al 2014; Ceinos, Guillot, Kelsh, Cerdá-Reveter, & Rotlland, 2015; Yong, Peichel, & McKinnon, 2015; Gu & Xia 2017; Kimura, Takehana, & Naruse, 2017; Roberts, Moore, & Kocher, 2017; Sefc et al, 2017; Lorin, Brunet, Laudet, & Volff, 2018; Kratochwil et al, 2018; Nagao et al, 2018; Cal et al, 2019; Lewis et al, 2019; Kon et al, 2020; Liang, Gerwin, Meyer, & Kratochwil, 2020). An increasing number of studies engaged fish research in high-throughput genomic approach of pigmentation variation (guppy: Tripathi et al, 2009; three-spine stickleback: Greenwood et al, 2011; Malek, Boughman, Dworkin, & Peichel, 2012; cichlids: O’Quin, Drilea, Conte, & Kocher, 2013; Henning, Jones, Franchini, & Meyer, 2013; Henning, Lee, Franchini, & Meyer, 2014; Albertson et al, 2014; Zhu et al, 2016; Roberts et al 2017; koi carp: Xu et al, 2014; arowana: Bian et al, 2016; goldfish: Kon et al, 2020).…”