Investigation of the effects of estrogen on skeletal gene expression during zebrafish larval head development

Richter²,

Sefc³

2017

Sci Rep

The diversity of fin morphology within and across fish taxa offers great, but still largely unexplored, opportunities to investigate the proximate mechanisms underlying fin shape variation. Relying on available genetic knowledge brought forth mainly by the comprehensive study of the zebrafish caudal fin, we explored candidate molecular mechanisms for the maintenance and formation of the conspicuously elongated filaments adorning the unpaired fins of the East African “princess cichlid” Neolamprologus brichardi. Via qPCR assays, we detected expression differences of candidate genes between elongated and short regions of intact and regenerating fins. The identified genes include skeletogenic and growth factors (igf2b, fgf3, bmp2 and bmp4), components of the WNT pathway (lef1, wnt5b and wnt10) and a regulatory network determining fin ray segment size and junction (cx43, esco2 and sema3d), as well as other genes with different roles (mmp9, msxb and pea3). Interestingly, some of these genes showed fin specific expression differences which are often neglected in studies of model fish that focus on the caudal fin. Moreover, while the observed expression patterns were generally consistent with zebrafish results, we also detected deviating expression correlations and gene functions.

Section: Methodsmentioning

confidence: 99%

A gene expression study of ornamental fin shape in Neolamprologus brichardi, an African cichlid species

Richter²,

Sefc³

2017

Sci Rep

“…GeNorm measures mean pairwise variation between each gene and other candidates, the expression stability or M value, and it excludes the gene with the highest M value (least stability) from subsequent analysis in a stepwise manner. NormFinder identifies the most stable genes (lowest expression stability values) based on analysis of the sample subgroups and estimation of inter- and intra-group variation in expression levels (Ahi et al, 2013; Pashay Ahi et al, 2016). …”

Section: Methodsmentioning

confidence: 99%

A gene expression study of dorso-ventrally restricted pigment pattern in adult fins ofNeolamprologus meeli, an African cichlid species

Sefc

2017

PeerJ

Fish color patterns are among the most diverse phenotypic traits found in the animal kingdom. Understanding the molecular and cellular mechanisms that control in chromatophore distribution and pigmentation underlying this diversity is a major goal in developmental and evolutionary biology, which has predominantly been pursued in the zebrafish model system. Here, we apply results from zebrafish work to study a naturally occurring color pattern phenotype in the fins of an African cichlid species from Lake Tanganyika. The cichlid fish Neolamprologus meeli displays a distinct dorsal color pattern, with black and white stripes along the edges of the dorsal fin and of the dorsal half of the caudal fin, corresponding with differences in melanophore density. To elucidate the molecular mechanisms controlling the differences in dorsal and ventral color patterning in the fins, we quantitatively assessed the expression of 15 candidate target genes involved in adult zebrafish pigmentation and stripe formation. For reference gene validation, we screened the expression stability of seven widely expressed genes across the investigated tissue samples and identified tbp as appropriate reference. Relative expression levels of the candidate target genes were compared between the dorsal, striped fin regions and the corresponding uniform, grey-colored regions in the anal and ventral caudal fin. Dorso-ventral expression differences, with elevated levels in both white and black stripes, were observed in two genes, the melanosome protein coding gene pmel and in igsf11, which affects melanophore adhesion, migration and survival. Next, we predicted potential shared upstream regulators of pmel and igsf11. Testing the expression patterns of six predicted transcriptions factors revealed dorso-ventral expression difference of irf1 and significant, negative expression correlation of irf1 with both pmel and igsf11. Based on these results, we propose pmel, igsf11 and irf1 as likely components of the genetic mechanism controlling distinct dorso-ventral color patterns in N. meeli fins.

“…To identify stably expressed reference genes for further analysis, we screened 6 genes which are known to be expressed in different tissues and often used as reference genes for qPCR analysis in fishes ( Table S1 ) ( Yang et al, 2013 ; Ahi et al, 2013 ; Pashay Ahi et al, 2016 ; Ahi & Sefc, 2017 ). Expression levels of the melanosome marker gene slc24a5 , which encodes an intracellular membrane cation exchanger predominantly present in melanosomes ( Lamason et al, 2005 ), and the iridophore lineage specific marker ltk, which encodes a tyrosine kinase receptor required for iridophore differentiation and stripe formation in adult zebrafish ( Lopes et al, 2008 ), were quantified as proxies for the densities of the two chromatophore types.…”

Section: Methodsmentioning

confidence: 99%

“…BestKeeper determines the stability of reference genes through a correlation calculation index (r). GeNorm estimates mean pairwise variation between each gene and other candidates ( M value) and NormFinder identifies the most stable genes (lowest expression stability values) through analysis of inter- and intra-group variation in expression levels ( Ahi et al, 2013 ; Pashay Ahi et al, 2016 ).…”

Section: Methodsmentioning

confidence: 99%

Anterior-posterior gene expression differences in three Lake Malawi cichlid fishes with variation in body stripe orientation

Sefc

2017

PeerJ

Morphological differentiation among closely related species provides opportunities to study mechanisms shaping natural phenotypic variation. Here, we address variation in the orientation of melanin-colored body stripes in three cichlid species of the tribe Haplochromini. Melanochromis auratus displays a common pattern of dark, straight horizontal body stripes, whereas in Aristochromis christyi and Buccochromis rhoadesii, oblique stripes extend from the anterior dorsal to the posterior mid-lateral trunk. We first validated a stably reference gene, and then, investigated the chromatophore distribution in the skin by assessing the expression levels of the iridophore and melanophore marker genes, ltk and slc24a5, respectively, as well as pmel, a melanophore pigmentation marker gene. We found anterior-posterior differences in the expression levels of the three genes in the oblique-striped species. The higher anterior expression of ltk, indicates increased iridophore density in the anterior region, i.e., uneven horizontal distribution of iridophores, which coincides with the anterior dorsalization of melanophore stripe in these species. The obliqueness of the horizontal body stripes might be a result of distinct migratory or patterning abilities of melanophores in anterior and posterior stripe regions which could be reflected by variation in the expression of genes involved in melanophore patterning. To address this, we investigated anterior-posterior expression levels of a primary set of candidate target genes with known functions in melanophore migration and stripe patterning in the adult zebrafish, and their related gene regulatory network. Among these genes, those with differences in anterior-posterior expression showed only species-specific differential expression, e.g., sdf1a, col14a1a, ifitm5, and agpat3, with the exception of fbxw4/hagoromo (differentially expressed in an oblique-and the straight-striped species). In summary, distinct anterior-posterior gradients in iridophore density found to be more similar characteristic between the two oblique-striped species. Furthermore, the species-specific differential expression of genes involved in stripe patterning might also implicate distinct molecular processes underlying the obliqueness of body stripe in two closely related cichlid species.