Gonzalo Riadi

The transition of vertebrates from aquatic to terrestrial environments represents one of the most profound evolutionary events in their history, involving extensive physiological and morphological innovations. Key adaptations included the transformation of fins into limbs with digits to enable efficient terrestrial locomotion, the ability to perceive novel environmental stimuli, and the emergence of reproductive strategies suited to life on land, processes in which ion channels played fundamental roles. Accordingly, understanding the genetic basis of vertebrate terrestrialization requires investigating the evolution of this group of membrane proteins. Our analyses reveal that the proportion of ion channel genes is highly conserved, representing approximately 1.4% to 1.6% of total protein-coding genes in most lineages, with a notable increase to ∼1.9% in teleost fishes. Our natural selection analyses revealed an overrepresentation of specific ion channel gene families, including TRP, RyR, HTR3, and HCN. We identified 29 ion channel genes showing signatures of positive selection, many of which are associated with key physiological functions such as nociception and thermosensation. We also detected an elevated rate of gene turnover in the common ancestor of terrestrial vertebrates, indicative of substantial genomic remodeling through gene gain and loss. Together, these findings suggest that, despite overall conservation in the proportions of ion channel genes, specific gene families underwent changes that were likely critical to meeting the physiological demands of terrestrial life. These results provide a foundation for future comparative and functional studies aimed at elucidating the molecular mechanisms underlying major environmental transitions. Significance: The transition from water to land was one of the most important events in vertebrate evolution, requiring animals to detect new environmental cues, sense temperature, coordinate movement, and cope with unfamiliar pathogens and physical stresses. Because ion channels control many of these physiological processes, they provide an ideal system for investigating the molecular basis of vertebrate terrestrialization. By tracing their evolutionary history across vertebrates, we show that many ion channels underlying these functions underwent adaptive evolution during the transition from water to land. Our findings suggest that adaptive changes in ion channels helped vertebrates acquire the physiological capabilities needed to survive and diversify on land, providing new insight into the molecular basis of one of the greatest phenotypic transformations in vertebrate evolution.

The eukaryotic genomes encode hundreds of proteins that function as ion channels and transporters. Essential for sustaining life, these proteins mediate the movement of inorganic ions (e.g., K ⁺ , Na ⁺ , Cl ⁻ , and Ca ²⁺ ) across the plasma membrane according to their electrochemical gradients. In multicellular organisms, a diverse array of ion channels contributes to the maintenance of the resting membrane potential, the regulation of pH, osmolarity, and cell volume, and the control of secretion, electrical excitability, and synaptic activity, among many other fundamental physiological processes. Although independent evolutionary origins have been proposed for several ion channel families, their relative hierarchical importance for cellular viability remains poorly understood. To advance our knowledge of ion channel evolutionary history, we focused on determining the minimal combination of permeabilities that allows cellular viability. To this end, we conducted a survey of representative prokaryotes with small genomes across bacterial and archaeal phyla. By focusing on the smallest genomes, our approach enabled the identification of five ion channel architectures shared among prokaryotes. Among these, non-selective mechanosensitive channels (MscS and MscL) are the most abundant, followed by CLC-type channels, RCK-containing 2TM potassium channels, and proton channels of the MotA/TolQ/ExbB family. The conservation of the mechanosensitive protein architecture across archaeal and bacterial membranes suggests that the capacity to monitor physical membrane integrity predates the requirements for electrical communication.

Aging has traditionally been studied through the lens of protein-coding genes, with a strong bias toward data derived from male organisms. As a result, the role of non-coding elements and potential sex-specific differences remains largely unexplored. Transposable elements (TEs), mobile sequences capable of altering genome structure and regulating gene expression, have recently gained attention for their roles in development and aging. However, despite this growing interest, key aspects of TE expression dynamics are still poorly characterized, particularly in female tissues. To address this gap, we analyzed TE expression in RNA-Seq liver tissue from male (8, 26, 60, 78, and 104 weeks) and female (3, 24, 48, and 72 weeks) mice. Our results reveal distinct TE expression trends between sexes. While previous studies report increased TE expression with aging, we identified a subset of TEs with decreasing expression over time, differing between males and females. We also observed inverse expression trends between a few TEs and their nearby genes, supporting a potential regulatory relationship. We identified TEs with changing expression (CE TEs) through age associated with nearby genes showing strong expression correlations (|ρ|≥ 0.6). In males, correlated genes such as Txnrd2, Mthfd1, and Dkk3 are involved in redox regulation, one-carbon metabolism, and Wnt signaling, respectively, while in females, Thrb and Cd55 are linked to metabolic regulation and immune protection. These associations suggest that TE activity may be functionally coupled to transcriptional programs relevant to liver physiology and aging. These findings highlight the importance of examining TE expression in both sexes and suggest their potential regulatory roles in age-related liver physiology.

TRPV1, a member of the transient receptor potential (TRP) family, is a non-selective cation channel primarily known for its role in pain perception, inflammation, and thermosensation. In mammals, it responds to noxious heat (>43°C) and chemical stimuli such as capsaicin and protons. It is widely expressed in sensory neurons, notably in the dorsal root and trigeminal ganglia. However, it is also found in some non-neuronal tissues, like the skin and bladder. The human canonical variant of TRPV1 renders a protein with 839 residues. Different splice variants have been described, and some display a dominant negative effect, partially or totally inhibiting the activity of the canonical counterpart. Here, we characterize a splice variant that encodes for a channel of 850 amino acids (TRPV1 ₈₅₀ ). This variant is an evolutionary novelty of catarrhine (Old World monkeys and apes) primates, incorporating an exon of 33 bp long. Both imaging of membrane expression and electrophysiological recordings suggest that TRPV1 ₈₅₀ alone does not reach the plasma membrane. However, in cells co-expressing the canonical and TRPV1 ₈₅₀ variants, the latter would act as a dominant negative, preventing the canonical variant from reaching the plasma membrane and rendering smaller macroscopic currents in response to capsaicin. Thus, this new isoform of the TRPV1 ion channel represents a novel form of functional regulation only present in apes and Old World monkeys. Significance: We identified a novel isoform of the TRPV1 ion channel, a protein essential for detecting pain and heat stimuli unique to apes and Old World monkeys. This isoform contains an additional exonic sequence encoding eleven amino acids, which originated in the common ancestor of apes and Old World monkeys. This additional exonic sequence disrupts a specific domain in the N-terminal region of the protein, resulting in functional differences. Unlike the canonical TRPV1 isoform, this variant cannot reach the plasma membrane independently. Instead, it exerts a dominant-negative effect by interfering with the canonical TRPV1 protein, reducing its ability to localize to the cell surface and diminishing its responsiveness to capsaicin, a well-known TRPV1 activator.

The differentiation of clones within grape cultivars, specifically Vitis vinifera, has significant potential for the wine industry. This differentiation involves associating morphological features or a genetic signature with a particular cultivar clone, which is a challenging task. It has been difficult to experimentally find genetic signatures that differentiate a pair of clones, despite evidence suggesting that genomic differences exist. Are there genetic or genomic differences in a pair of clones? If so, where are the variations in the genome? Are there variations in protein coding genes? We addressed these questions by performing a bioinformatic analysis to identify genetic differences between certified clones of the same cultivar. Utilising genome sequencing data from tissue samples, we identified genomic positions differing between the clones and their cultivar reference genome, meeting the filtering criteria. Applying this approach to the Carménère and Merlot clones resulted in 5718 and 5218 variations, respectively, that differentiated the clones. Visual validation of 50 variations per cultivar revealed that 12% of these variations were located in the Merlot genes, while 32% were found in the Carménère genes. We estimated between 600 and 1000 variations per cultivar that could be validated by visual inspection. Despite the presence of these variations within genes, none was found to have a disruptive effect on protein function. By comparing our results with those of previous studies, we discuss issues pertaining to clone differentiation. In conclusion, there are genomic variations in pairs of clones that allow for their differentiation, though the variations are not directly related to the phenotype.

Cetaceans represent a natural experiment within the tree of life in which a lineage changed from terrestrial to aquatic habitats. This shift involved phenotypic modifications, representing an opportunity to explore the genetic bases of phenotypic diversity. Among the different molecular systems that maintain cellular homeostasis, ion channels are crucial for the proper physiological functioning of all living species. This study aims to explore the evolution of ion channels during the evolutionary history of cetaceans. To do so, we created a bioinformatic pipeline to annotate the repertoire of ion channels in the genome of the species included in our sampling. Our main results show that cetaceans have, on average, fewer protein-coding genes and a higher percentage of annotated ion channels than non-cetacean mammals. Signals of positive selection were detected in ion channels related to the heart, locomotion, visual and neurological phenotypes. Interestingly, we predict that the NaV1.5 ion channel of most toothed whales (odontocetes) is sensitive to tetrodotoxin, similar to NaV1.7, given the presence of tyrosine instead of cysteine, in a specific position of the ion channel. Finally, the gene turnover rate of the cetacean crown group is more than three times faster than that of non-cetacean mammals.

Cetaceans could be seen as a natural experiment within the tree of life in which a mammalian lineage changed from terrestrial to aquatic habitats. This shift involved extensive phenotypic modifications, which represent an opportunity to explore the genetic bases of phenotypic diversity. Furthermore, the availability of whole genome sequences in representative species of all main cetacean groups means that we are in a golden age for such studies. Among the different molecular systems that maintain cellular homeostasis, ion channels are crucial for the proper physiological functioning of all living species. This study aims to explore the evolution of ion channels during the evolutionary history of cetaceans. To do so, we created a bioinformatic pipeline to annotate the repertoire of ion channels in the genome of the species included in our sampling. Our main results show that cetaceans have on average, fewer protein-coding genes and a higher percentage of annotated ion channels than non-cetacean mammals. Signals of positive selection were detected in ion channels related to the heart, locomotion, visual and neurological phenotypes. Interestingly, we predict that the Na _V 1.5 ion channel of most toothed whales (odontocetes) is sensitive to tetrodotoxin (TTX), similar to Na _V 1.7, given the presence of tyrosine instead of cysteine, in a specific position of the ion channel. Finally, the gene turnover rate of the cetacean crown group is more than three times faster than non-cetacean mammals.

The embryo-maternal interaction occurs during the early stages of embryo development and is essential for the implantation and full-term development of the embryo. In bovines, the secretion of interferon Tau (IFNT) during elongation is the main signal for pregnancy recognition, but its expression starts around the blastocyst stage. Embryos release extracellular vesicles (EVs) as an alternative mechanism of embryo-maternal communication. The aim of the study was to determine whether EVs secreted by bovine embryos during blastulation (D5-D7) could induce transcriptomic modifications, activating IFNT signaling in endometrial cells. Additionally, it aims to assess whether the EVs secreted by embryos produced in vivo (EVs-IVV) or in vitro (EVs-IVP) have different effects on the transcriptomic profiles of the endometrial cells. In vitro- and in vivo-produced bovine morulae were selected and individually cultured for 48 h to collect embryonic EVs (E-EVs) secreted during blastulation. E-EVs stained with PKH67 were added to in vitro-cultured bovine endometrial cells to assess EV internalization. The effect of EVs on the transcriptomic profile of endometrial cells was determined by RNA sequencing. EVs from both types of embryos induced several classical and non-classical IFNT-stimulated genes (ISGs) and other pathways related to endometrial function in epithelial endometrial cells. Higher numbers of differentially expressed genes (3552) were induced by EVs released by IVP embryos compared to EVs from IVV (1838). Gene ontology analysis showed that EVs-IVP/IVV induced the upregulation of the extracellular exosome pathway, the cellular response to stimulus, and the protein modification processes. This work provides evidence regarding the effect of embryo origin (in vivo or in vitro) on the early embryo-maternal interaction mediated by extracellular vesicles.