Biology subjects

Gentile, I.

Publications and source records attributed to Gentile, I..

6 recordsLinked to original sources

bioRxiv2025

Pan-angiosperm analysis of the CLE signaling peptide family unveils paths, patterns, and predictions of paralog diversification

The compositions of conserved gene families often vary widely between species, complicating predictions and experimental tests of shared versus distinct functions, especially in families shaped by extensive duplication, redundancy, and paralog diversification. The plant CLV3/EMBRYO- SURROUNDING REGION (CLE) small-signaling peptide family exemplifies these challenges. Although genetic studies in model systems have identified shared roles for a few CLE genes and species-specific redundancies, an evolutionary analysis of the entire family over deep time could empower predictive and experimental dissections of functions obscured by redundancy. We developed a scanning pipeline that de novo annotated CLE genes from 2,000 genomes representing 1,000 species, uncovering thousands of previously undetected family members and producing a comprehensive phylogenetic reconstruction and tracing of the familys evolution and sequence diversification over 140 million years. Computational modeling of coding and cis-regulatory regions predicted lineage-specific asymmetries in paralog redundancy, stemming from ancestral amino acids in the functional core of the dodecapeptide and partial conservation of promoter elements. We tested these predictions using two genome-editing strategies in Solanaceae. Base- editing of deeply conserved residues in the CLV3 dodecapeptide and its paralogs across three species confirmed their critical roles in repressing stem-cell proliferation, and multiplex CRISPR knockouts of the 52 tomato CLE genes resolved pairwise and higher-order redundancies, revealing previously uncharacterized regulators of shoot architecture and plant size. These findings show how both peptide and cis-regulatory erosion shape CLE redundancy and provide a framework for detecting and translating deep evolutionary signals into testable genetic hypotheses across compositionally complex gene families.

plant biology↗

bioRxiv2025

Effect of plant tissue culture parameters on the ploidy level of Physalis grisea, Solanum lycopersicum, and Solanum prinophyllum regenerants

Plants regenerated from seedling explants (hypocotyls and cotyledons) of the Solanaceae family members Physalis grisea (groundcherry), Solanum lycopersicum (tomato), and Solanum prinophyllum (forest nightshade) were used to determine the in vitro culture parameters that contribute to the incidence in polyploidization of tissue culture-derived plants (regenerants) from these species. We examined the possible effects of zeatin concentration in the plant regeneration medium, explant source, and species. Plants were grown to maturity under greenhouse conditions, pollen was collected and germinated. Flow cytometry analysis verified the utility of the pollen germination method for determining differences in ploidy, which was based on the number of pollen tubes produced with one tube representing diploid and two indicating polyploid. As for zeatin concentration, we assessed the effect of our standard method of initiation on medium containing 2 mg/l followed by 1 mg/l 2 weeks after culture initiation in comparison with 0.25, 0.5, and 1 mg/l throughout the culture lifetime. There were no major correlations for zeatin concentration on ploidy status across the species except for plants regenerated from S. lycopersicum hypocotyl explants where the percentage of polyploid regenerants increased with increasing concentrations. As for species and explant effects, P. grisea plants regenerated from hypocotyl explants had the highest percentage of polyploid plants at 81% compared to 43% and 35% for S. lycopersicum and S. prinophyllum, respectively. From cotyledons, 8% of S. lycopersicum and 20% of S. prinophyllum were polyploid. A comparison with P. grisea could not be made because cotyledon explants do not regenerate on zeatin-containing medium. The results indicated the incidence of polyploidization cannot be generalized for zeatin concentration, however, an influence of explant type and species was observed. Effects of increased ploidy on plant morphology were primarily larger flower and seed size; however, no significant differences were observed in plant or fruit size.

plant biology↗

bioRxiv2024

Solanum pan-genomics and pan-genetics reveal paralogs as contingencies in crop engineering

Pan-genomics and genome editing technologies are revolutionizing the breeding of globally cultivated crops. A transformative opportunity lies in the reciprocal exchange of genotype-to-phenotype knowledge of agricultural traits between these major crops and hundreds of locally cultivated indigenous crops, thereby enhancing the diversity and resilience of our food system. However, species-specific genetic variants and their interactions with desired natural or engineered mutations pose barriers to achieving predictable phenotypic effects, even between closely related crops or genotypes. Here, by establishing a pan-genome of the crop-rich genus Solanum and integrating functional genomics and genetics, we show that gene duplication and subsequent paralog diversification are a major obstacle to genotype-phenotype predictability. Despite broad conservation of gene macrosynteny among chromosome-scale references for 22 species, including 13 indigenous crops, hundreds of global and lineage-specific gene duplications exhibited dynamic evolutionary trajectories in paralog sequence, expression, and function, including among members of key domestication gene families. Extending our pan-genome with 10 cultivars of African eggplant and leveraging quantitative genetics and genome editing, we uncovered an intricate history of paralog emergence and evolution within this indigenous crop. The loss of an ancient redundant paralog of the classical regulator of stem cell proliferation and fruit organ number, CLAVATA3 (CLV3), was compensated by a lineage-specific tandem duplication. Subsequent pseudogenization of the derived copy followed by a cultivar-specific structural variant resulted in a single fused functional copy of CLV3 that modifies locule number alongside a newly identified gene controlling the same trait. Our findings demonstrate that paralog diversifications over short evolutionary periods are critical yet underexplored contingencies in trait evolvability and independent crop domestication histories. Unraveling these contingencies is crucial for translating genotype-to-phenotype relationships across related species.

genetics↗

bioRxiv2024

The sodium-proton exchangers sNHE and NHE1 control plasma membrane hyperpolarization in mouse sperm.

Sperm capacitation, crucial for fertilization, occurs in the female reproductive tract and can be replicated in vitro using a medium rich in bicarbonate, calcium, and albumin. These components trigger the cAMP-PKA signaling cascade, proposed to promote hyperpolarization of the mouse sperm plasma membrane through activation of SLO3 K+ channel. Hyperpolarization is a hallmark of capacitation: proper membrane hyperpolarization renders higher in vitro fertilizing ability, while Slo3 KO mice are infertile. However, the precise regulation of SLO3 opening remains elusive. Our study challenges the involvement of PKA in this event and reveals the role of Na+/H+ exchangers. During capacitation, calcium increase through CatSper channels activates NHE1, while cAMP directly stimulates the sperm-specific NHE, collectively promoting the alkalinization threshold needed for SLO3 opening. Hyperpolarization then feeds back Na+/H+ activity. Our work is supported by pharmacology, and a plethora of KO mouse models, and proposes a novel pathway leading to hyperpolarization. TeaserAlkalinization of sperm cytoplasm activates potassium channels to hyperpolarize the plasma membrane in a PKA independent cascade.

cell biology↗

bioRxiv2024

Convergent evolution of plant prickles is drivenby repeated gene co-option over deep time

An enduring question in evolutionary biology concerns the degree to which episodes of convergent trait evolution depend on the same genetic programs, particularly over long timescales. Here we genetically dissected repeated origins and losses of prickles, sharp epidermal projections, that convergently evolved in numerous plant lineages. Mutations in a cytokinin hormone biosynthetic gene caused at least 16 independent losses of prickles in eggplants and wild relatives in the genus Solanum. Strikingly, homologs promote prickle formation across angiosperms that collectively diverged over 150 million years ago. By developing new Solanum genetic systems, we leveraged this discovery to eliminate prickles in a wild species and an indigenously foraged berry. Our findings implicate a shared hormone-activation genetic program underlying evolutionarily widespread and recurrent instances of plant morphological innovation.

evolutionary biology↗

bioRxiv2022

Dynamic evolution of small signaling peptide compensation in plant stem cell control

Gene duplications are a hallmark of plant genome evolution and a foundation for genetic interactions that shape phenotypic diversity1-5. Compensation is a major form of paralog interaction6-8, but how compensation relationships change as allelic variation accumulates is unknown. Here, we leveraged genomics and genome editing across the Solanaceae family to capture the evolution of compensating paralogs. Mutations in the stem cell regulator CLV3 cause floral organs to overproliferate in many plants9-11. In tomato, this phenotype is partially suppressed by transcriptional upregulation of a closely related paralog12. Tobacco lost this paralog, resulting in no compensation and extreme clv3 phenotypes. Strikingly, the paralogs of petunia and groundcherry nearly completely suppress clv3, indicating a potent ancestral state of compensation. Cross-species transgenic complementation analyses show this potent compensation partially degenerated in tomato due to a single amino acid change in the paralog and cis-regulatory variation that limits its transcriptional upregulation. Our findings show how genetic interactions are remodeled following duplications, and suggest that dynamic paralog evolution is widespread over short time scales and impacts phenotypic variation from natural and engineered mutations.

plant biology↗