The Chromatin Assembly Factor 1 is a heterotrimeric complex responsible for the nucleosome assembly during DNA replication and DNA repair. In humans, the largest subunit P150 is the major actor of this process. It has been recently considered as a tumor-associated protein due to its overexpression in many malignancies. Structural and functional studies targeting P150 are still limited and only scarce information about this subunit is currently available. Literature data and bioinformatics analysis assisted the identification of a stable DNA binding domain, encompassing residues from 721 to 860 of P150 within the full-length protein. This domain was recombinantly produced and in vitro investigated. An acidic region modulating its DNA binding ability was also identified and characterized. Results showed similarities and differences between the P150 and its yeast homologue, namely Cac-1, suggesting that, although sharing a common biological function, the two proteins may also possess different features.

Systemin (Sys) is an octadecapeptide, which upon wounding, is released from the carboxy terminus of its precursor, Prosystemin (ProSys), to promote plant defenses. Recent findings on the disordered structure of ProSys prompted us to investigate a putative biological role of the whole precursor deprived of the Sys peptide. We produced transgenic tomato plants expressing a truncated ProSys gene in which the exon coding for Sys was removed and compared their defense response with that induced by the exogenous application of the recombinant truncated ProSys (ProSys_(1-178), the Prosystemin sequence devoid of Sys region). By combining protein structure analyses, transcriptomic analysis, gene expression profiling and bioassays with different pests, we demonstrate that truncated ProSys promotes defense barriers in tomato plants through a hormone-independent defense pathway, likely associated with the production of oligogalacturonides (OGs). Both transgenic and plants treated with the recombinant protein showed the modulation of the expression of genes linked with defense responses and resulted in protection against the lepidopteran pest Spodoptera littoralis and the fungus Botrytis cinerea. Our results suggest that the overall function of the wild-type ProSys is more complex than previously shown, as it might activate at least two tomato defense pathways: the well-known Sys-dependent pathway connected with the induction of jasmonic acid biosynthesis and the successive activation of a set of defense-related genes, and the ProSys_(1-178)-dependent pathway associated with OGs production leading to the OGs mediate plant immunity.

In an increasingly challenging agricultural environment, the identification of novel tools for protecting crops from stress agents while securing marketable production is a key objective. Here we investigated the effects of three previously characterized Prosystemin-derived functional peptide fragments as protective agents against salt stress and as biostimulants modulating tomato yield and quality traits. The treatments of tomato plants with femtomolar amounts of the peptides alleviated salt stress symptoms, likely due to an increase in root biomass up to 18% and the upregulation of key antioxidant genes such as APX2 and HSP90. In addition, the peptides exhibited biostimulant activity, significantly improving root area (up to 10%) and shoot growth (up to 9%). We validated such activities through two-year field trials carried out on industrial tomato crops. Peptide treatments confirmed their biostimulant effects, leading to a nearly 50% increase in marketable production compared to a commonly used commercial product and consistently enhancing fruit °Brix values.

Plants constantly face multiple biotic and abiotic stresses, which they counteract through sophisticated immune mechanisms based on the recognition of endogenous and exogenous elicitors. Tomato Systemin (Sys), released from the C-terminal region of Prosystemin (ProSys), is a well-known peptide that activates jasmonic acid (JA)-dependent defense responses. In contrast, the N-terminal fragment ProSys_(1-178), lacking the Sys peptide, has recently emerged as an independent defense inducer, likely involving an OG-mediated pathway distinct from the JA signalling triggered by Sys. In this study, we investigated the molecular and biological responses elicited by ProSys_(1-178) in comparison with Sys by combining comparative transcriptomic analyses of ProSys_(1-178) and full-length ProSys (ProSys_FL) transgenic tomato plants with bioassays of peptide treated plants. Transcriptomic analysis revealed that ProSys_(1-178) induced a substantially broader transcriptional reprogramming than ProSys_FL, with only 46 differentially expressed genes in common, that displayed opposite expression patterns. Both transgenic lines conferred resistance against Spodoptera littoralis and Botrytis cinerea. Moreover, exogenous application of ProSys_(1-178) and Sys differentially modulated local and systemic defense-related gene expression and significantly reduced B. cinerea disease severity. Although Sys provided stronger protection against the fungal pathogen, ProSys_(1-178) modulated a broader set of defense-related genes, suggesting the activation of a distinct immune pathway. The induction of a polygalacturonase gene, together with the previous detection of short OGs in ProSys_(1-178)-expressing plants, is consistent with, but does not establish, an OG-associated component of this response. Overall, our findings support a multifaceted role for ProSys_(1-178) in tomato immunity.

Sola l 4.02 is a major tomato allergen of the PR-10 family and a Bet v 1 homolog implicated in pollen–food cross-reactivity, yet its molecular properties remain poorly defined. Here, we report the recombinant production and its comprehensive structural and functional characterization. Sequence and structural analysis support a canonical Bet v 1-like fold including the conserved hydrophobic ligand binding site cavity typical of PR-10 proteins. Biophysical analyses showed that recombinant Sola l 4.02 is a compact, monomeric, and well-folded protein in solution. Circular dichroism measurements revealed moderate thermal stability, with a melting temperature of 57.5 °C and partial recovery of the native conformation after cooling, indicating partially reversible unfolding. The protein remained structurally stable between pH 5.5 and 9.0, whereas mildly acidic conditions promoted partial destabilization. To probe ligand recognition, we tested selected phenolic compounds by microscale thermophoresis. Sola l 4.02 bound quercetin-3-glucoside, naringenin, and daidzein with low micromolar affinities, while caffeic acid showed only weak interaction and hydroxycinnamic acid displayed no detectable binding. Limited proteolysis assays further demonstrated that ligand binding modulates protein susceptibility to digestion. In the presence of bound ligands, Sola l 4.02 displayed enhanced resistance to both trypsin and α-chymotrypsin, with naringenin and daidzein providing the strongest effect. These results identify Sola l 4.02 as a folded, ligand-binding PR-10 allergen whose proteolytic stability is influenced by small-molecule association, providing new insights into the relationship between structural dynamics, ligand occupancy, and the molecular behavior of tomato food allergens.