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Fuentes, R.

Publications and source records attributed to Fuentes, R..

7 recordsLinked to original sources
bioRxiv2025

The olfactory organ is a site for neuroendocrine modulation of reproduction in zebrafish

Gonadotropin-releasing hormone (GnRH) is one of the most fascinating neuroendocrine peptides: It is essential for maintaining the reproductive state of vertebrates and also displays high sequence homology to the tridecapeptide mating pheromone of the yeast, S. cerevisiae. In spite of its highly conserved role in vertebrate reproduction, recent studies in zebrafish show that the loss of function of genes encoding Gnrh isoforms does not cause infertility. Here we first investigated whether Phoenixin, a novel peptide acting in the reproductive pathway of vertebrates, is the hormone that has replaced Gnrh in zebrafish. While loss of function of the phoenixin gene affected female differentiation, we observed no defects in fertility. We next reconsidered the GnRH pathway and turned to the natural world, where fishes use waterborne hormones to control reproduction. Thus, we investigated whether exogenous Gnrh affects the hypothalamic-pituitary axis. Here we show that fish isolated from their conspecifics and kept in artificial water devoid of fish odors, mounted sex-appropriate pituitary responses when Gnrh was added to the water. Furthermore, blocking the access of water to the olfactory organs eliminated these responses. We then analyzed Gnrh signaling by knocking out the gene encoding gnrh-receptor3 (gnrh-r3) and, surprisingly, found that fish homozygous for a gnrh-r3 null mutation were almost completely infertile: males did not produce sperm and females produced but a few mature oocytes. Finally, we found that Gnrh was present in nanomolar concentrations in the water that houses the fish, thus supporting the hypothesis that waterborne Gnrh from conspecifics plays a key role in regulating zebrafish reproduction in the absence of the endogenous ligand.

neuroscience↗
bioRxiv2025

Zebrafish armc9 mutant reveals a link between ARM-Type fold domain loss, inflammation, and spinal curvature

The vertebral column (VC) defines the central axis of vertebrates. Among its malformations, adolescent idiopathic scoliosis (AIS) is the most common, characterized by lateral spine curvature without congenital abnormalities. While its etiology remains elusive, genetic studies in zebrafish have linked AIS to skeletogenesis, cilia structure, cerebrospinal fluid flow, and inflammation. In this study, we characterized the zebrafish armc9 mutant, exhibiting AIS-like curvature without vertebral malformations, subcommissural organ, or Reissner fiber structure disruptions. We also detected an inflammatory response at the curvature origin. Structural bioinformatics and molecular docking analyses suggest that the mutant Armc9 protein partially lacks the functional ARM-type fold domain, which is crucial for its interaction with Togaram1, revealing that both proteins are relevant for ciliary signaling and spine curvature. Phylogenetic analyses indicate that the ARMC gene family encompasses 12 members, highlighting that the Armc9 gene has deep evolutionary roots. Our results demonstrate that the zebrafish armc9 mutant accurately represents the AIS phenotype, suggesting that the loss of Armc9s C-terminal ARM-type fold domain could affect the ciliary integrity and contribute to spine curvature by altering the skeletogenic environment. Taken together, this study proposes that the zebrafish armc9 mutant offers a unique opportunity to understand the etiology of AIS and provides new insights into the molecular interaction networks of Armc9 in ciliary signaling.

developmental biology↗
bioRxiv2023

Behavioral Thermoregulation in Captive Fish: Molecular, Physiological, and Welfare Implications

Environmental Enrichment (EE) serves as a cornerstone in the attempt to emulate natural habitats for captive organisms. While substantial strides have been made in this field, current methodologies still grapple with discrepancies between recreated habitats and the innate conditions vital for maintaining biological homeostasis in captive species. Our study highlights the pivotal role of behavioral thermoregulation in modulating molecular and physiological outcomes in captive fish. Collective evidence suggests that enabling fish to autonomously regulate temperature confers numerous beneficial cellular and systemic effects. Specifically, introducing a thermal gradient within the EE paradigm correlated with increased survival metrics, enhanced physiological parameters, and improved welfare indices, establishing the criticality of thermoregulation in captivity. In contrast, the lack of a thermoregulatory framework resulted in the emergence of transient free radicals, a clear marker of temperature-induced oxidative stress. Persistent disruptions in free radical equilibrium, especially in uniform temperature settings, were linked to DNA damage, heightened cellular apoptosis, tissue anomalies, and metabolic deviations. In conclusion, this research underscores the significance of behavioral thermoregulation as an integral feature of EE, especially related to fish in controlled environments. Our data present key biomarkers valuable for optimizing fish welfare and highlight the necessity for sustained research into their adaptability and survival benchmarks. Such insights aim to enhance EE protocols, fortifying their efficacy in mirroring natural habitats and, in turn, advancing the welfare benchmarks of captive organisms.

physiology↗
bioRxiv2023

Maternal regulation of the vertebrate egg-to-embryo transition

Egg activation and early embryonic development are complex and highly regulated processes that involve a series of coordinated cellular and molecular events after fertilization. While significant progress has been made in unraveling the mechanisms underlying these processes, there remains a need to comprehensively understand the precise molecular pathways and regulatory factors involved. We characterized four recessive maternal-effect mutants identified from a zebrafish forward genetic screen that function during the egg-to-embryo transition. We found that these genes encompass distinct aspects of egg activation, including cortical granule biology, cytoplasmic segregation, and the suppression of microtubule organizing center (MTOC) assembly and ectopic aster-like microtubule formation. These genes are essential to the development of the early embryo and the establishment of the basic body plan. Notably, we discovered a novel gene that we named krang, which is highly conserved across metazoans. Maternal Krang was found to be associated with the function of cortical granules during egg activation. This collection of mutants represents valuable tools to understand the genetic architecture underlying phenotypic traits shaping the egg-to-embryo transition. Furthermore, these results highlight the evolutionary conservation of maternal functions in diverse species. By deepening our understanding of these findings, we will improve our knowledge of reproductive traits and potentially develop new diagnostic tools to address human reproductive disorders. Author SummaryDuring egg activation, the egg undergoes a series of biochemical and cellular modifications that activate its dormant metabolism, preparing it to initiate embryonic development. Egg activation initiates the completion of meiosis, and the newly formed zygote undergoes a series of additional changes, including the cell cycle establishment. However, our knowledge of the molecular mechanisms controlling these processes remain limited. We identified four recessive maternal-effect mutants in zebrafish that exhibit a range of developmental alterations during egg activation and early embryogenesis. We found that these maternally acting mutant genes are associated with defects in cortical granule exocytosis, yolk-cytoplasm segregation, microtubule nucleation and dynamics that are critical for the egg-to-embryo transition. Our results suggest that the proper regulation of these processes is essential for successful egg development and embryogenesis. We characterized the Krang factor, which regulates aspects of egg activation likely by modulating the secretory pathway. Further studies on this gene may provide new insights into the molecular mechanisms underlying oocyte development and egg quality acquisition. Overall, our collection of maternal-effect mutants sheds light on the proper regulation of key molecular and cellular events for successful egg development, with important implications for reproductive medicine and assisted reproductive technologies.

developmental biology↗
bioRxiv2023

Maternal regulation of the vertebrate oocyte-to-egg transition

Maternally-loaded factors in the egg accumulate during oogenesis and are essential for the oocyte and egg to acquire developmental competence and ensure the production of viable embryos. The oocyte-to-egg transition consists of the regulation of multiple molecular processes both cytoplasmic and nuclear acting in the late oocyte during a process called oocyte maturation. However, the molecular nature and functional importance of factors acting at this stage remain poorly understood. Here, we present a collection of 5 recessive maternal-effect mutants identified in a zebrafish forward genetic screen that reveal unique molecular insights into the mechanisms controlling the vertebrate oviparous oocyte-to-egg transition. We identified critical cytoplasmic regulators of yolk globule formation and maturation that are essential for egg development and embryogenesis. Specifically, the maternal-effect genes, over easy, poached, p33bjta, and black caviar control yolk globule sizing and/or protein cleavage during oogenesis, likely through endolysosomal organization independent of nuclear oocyte maturation. Furthermore, we cloned one of the mutant genes, identifying a subunit of the Adaptor Protein complex 5, which regulates intracellular trafficking, and yolk vesicle formation. Together, these mutant genes represent novel genetic entry points to decipher the molecular mechanisms functioning in the oocyte-to-egg transition, fertility, and human disease. Additionally, our genetic screen provides valuable functional tools for exploring the evolutionary fates of maternal factors and their contribution to developmental strategies for reproductive success in metazoans. Author SummaryThe oocyte-to-egg transition consists of the coordinated regulation of multiple molecular processes acting in the late oocyte. This transcriptionally silent period requires the precisely timed function of maternally-supplied gene products during oogenesis. However, knowledge of their molecular nature and in vivo function remains incomplete. The mutants reported here provide access to maternal factors regulating the processes that prepare an oocyte for reproductive competence and embryogenesis. We have identified essential regulators of yolk granule formation and protein processing. Specifically, we found that the highly conserved maternal Ap5m1 protein regulates yolk granule maturation, which generate essential nutrients and immunity for growth and development in oviparous animals. The mutants presented here represent attractive genetic models to investigate the molecular and cell biological mechanisms that control the oocyte-to-egg transition, as well as reveal a collection of genetic factors indispensable for reproduction and survival. Importantly, knowledge of their genetic underpinnings and biological importance in reproduction will also pave the way to investigate the evolution of maternal genes during vertebrate development.

developmental biology↗
bioRxiv2018

The Hippo pathway effector Taz is required for cell fate specification and fertilization in zebrafish

In the last decade, Hippo signaling has emerged as a critical pathway integrating extrinsic and intrinsic mechanical cues to regulate cell proliferation and survival, tissue morphology and organ size in vivo. Despite its essential role in organogenesis, surprisingly much less is known about how it connects biomechanical signals to control of cell fate and cell size during development. Here we unravel a novel and unexpected role of the Hippo pathway effector Taz (wwtr1) in the control of cell size and cell fate specification. In teleosts, fertilization occurs through a specific structure at the animal pole, called the micropyle. This opening in the chorion is formed during oogenesis by a specialized somatic follicle cell, the micropylar cell (MC). The MC has a peculiar shape and is much larger than its neighboring follicle cells but the mechanisms underlying its specification and cell shape acquisition are not known. Here we show that Taz is essential for the specification of the MC and subsequent micropyle formation in zebrafish. We identify Taz as the first bona fide MC marker and show that Taz is specifically and strongly enriched in the MC precursor before the cell can be identified morphologically. Altogether, our genetic data and molecular characterization of the MC lead us to propose that Taz is a key regulator of the MC fate activated by physical cues emanating from the oocyte to initiate the MC morphogenetic program. We describe here for the first time the mechanism underlying the specification of the MC fate.

developmental biology↗