UNCOVERING THE EPIGENETIC LANDSCAPE OF LARVAL ONTOGENY IN Sparus aurata – PROSPECTS FOR SHAPING DEVELOPMENT

Introduction

Considerable efforts have been devoted to developing protocols aimed at driving farmed fish towards desired phenotypes. However, success has remained limited, likely because phenotypic programming is largely mediated through epigenetic mechanisms, while current knowledge on the epigenetic regulators (EpiRegs) that ultimately transfer environmental information to the genome remains scarce. Thus, there is a need to better understand the bases of this phenotypic plasticity to advance toward phenotypes that fit future aquaculture conditions. The main objectives of the present study were, first, to develop and validate a qPCR array for EpiRegs in gilthead seabream (Sparus aurata) and, second, to apply this tool to characterize, for the first time, the ontogenetic changes in EpiRegs during larval development of this species.

Materials and Methods

A qPCR array was specifically developed for Sparus aurata to profile epigenetic regulators. The design included 73 target genes identified as EpiRegs, followed by technical validation. The array was assembled in 384-well microplates and encompassed genes involved in key regulatory processes, including enzymes, enzyme regulators, transcription factors, transcription factor regulators, and methyl-binding proteins. Gene expression dynamics were assessed across four ontogenetic stages: before hatching, hatching, 6 days post-hatching (dph), and 20 dph. Besides, to assess gilthead seabream EpiRegs susceptibility to manpulation during early development, an additional experiment was conducted in which larvae were fed microdiets supplemented with an epigenetic modulator, and sampled at the same developmental stages.

Results

Initial results revealed that EpiRegs displayed ontogenetic expression patterns throughout larval development in gilthead seabream. Hierarchical clustering analysis identified several groups of genes sharing similar temporal dynamics across developmental stages (Fig. 1). While some EpiRegs showed high expression levels during hatching followed by a marked downregulation at later stages, others exhibited the opposite trend, with progressive activation during post-hatch development. Several key EpiRegs, including those involved in DNA methylation (e.g., dnmt3ba, dnmt3l, and uhrf1), chromatin remodeling and transcriptional repression (e.g., mbd2), growth regulation (e.g., myca), and DNA damage response (e.g., gadd45b), exhibited ontogenetic expression patterns during larval development. Furthermore, dietary supplementation with an epigenetic modulator altered the expression dynamics of several EpiRegs, modifying their natural ontogenetic profiles, indicating that gilthead seabream epigenome may be susceptible to nutritional modulation during critical developmental windows.

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Figure 1. Hierarchical clustering analysis of EpiRegs expression profiles across larval developmental stages (B-HT: behore hatching; HT: hatching; 6 and 20 days post-hatching) in gilthead seabream (Sparus aurata).

Discussion

These results support the existence of a coordinated epigenetic regulatory program during early development in gilthead seabream. The ontogenetic variations observed in several EpiRegs highlights the dynamic nature of epigenetic regulation during critical developmental windows. Moreover, the modulation of these expression patterns by a dietary compound suggests that this regulatory program is susceptible to external nutritional cues in seabream. Ongoing analyses are currently addressing the relationship between EpiRegs expression patterns and tissue-specific changes in DNA methylation during development.

Together, these findings provide new insight into the molecular basis of developmental programming in fish and identify EpiRegs as promising targets for future nutriepigenomic strategies for presision nutrition in aquaculture.

Acknowledgment

This research was funded by EpiTool4Fish Project (PCM_00111) from the Complementary Plan for Marine Sciences ThinkInAzul Andalucía. C.N-G is supported by a Ramón y Cajal contract (RYC2024-050256-I) funded by MICIU, AEI and FSE+.