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Figure 1. 

Gene Ontology (GO) classification of proteins identified in the seed coat tissue of common bean. (a) Distribution of GO terms by ontology, showing the proportion of proteins assigned to the categories: Cellular Component, Molecular Function, and Biological Process. (b) Top five GO terms within each ontology ranked by frequency of annotation for each category.

Figure 2. 

Volcano plot showing the number of up- and down-regulated proteins (q-value < 0.05) in the seed coat tissue of common bean between day 0 and 4 after seed imbibition. Each point represents one protein, plotted according to its log₂FC change (x-axis) and –log₁₀ (q-value) (y-axis). Proteins with q-value ≥ 0.05 were classified as not significant (NS, gray). Among significant proteins (q-value < 0.05), those with positive log₂FC were considered upregulated (green), while those with negative log₂FC were considered downregulated (red). The numbers shown in the legend correspond to the count of proteins in each category. Proteomic profiles were compared using samples from four independent biological pools for each condition.

Figure 3. 

Top 40 Gene Ontology (GO) functional categories showing changes in protein abundance in the seed coat of common bean between day 0 and 4 of germination. % change values represent the percentage change in protein abundance between the two times. Bars represent the summed % change for all proteins annotated to each GO term, with positive values (green) indicating overall upregulation and negative values (red) indicating overall downregulation. GO annotations were obtained from the Gene Ontology database (http://geneontology.org). Only categories from the Molecular Function (MF) and Biological Process (BP) domains were included. Proteins with significant change (q-value < 0.05) were retained, and the categories were ranked by total positive % change; the top 40 terms are shown.

Figure 4. 

Top 40 KOG categories showing increases in protein abundance in the seed coat of common bean vulgaris between day 0 and 4 after imbibition. Only proteins with positive % change were retained. When proteins were annotated with multiple KOG terms, annotations were split and evaluated pairwise with their descriptions; entries lacking a description or labeled as uncharacterized/NA were excluded. Bars represent the summed % change across all proteins assigned to each KOG description (x-axis). Categories on the y-axis are ordered by the number of contributing proteins (most proteins at the top), while bar length reflects the total % change for that category. KOG annotations were obtained from the NCBI COG/KOG resource. Only significant change proteins with a q-value < 0.05 were retained for analysis.

Figure 5. 

Expression pattern of (a) Ribonucleases T2, (b) nucleoside diphosphate kinase, and (c) nucleases in seed coats at 0 (0 D) and 4 (4 D) d after the start of imbibition. Gene expression analysis was performed using qRT-PCR on total RNA samples extracted from the seed coat of common bean. The relative expression level was normalized using the geometric mean of two reference genes and analysed using the 2^−ΔCᴛ method. Student´s t-test was performed comparing both conditions. Statistical significance levels were indicated as follows: * p < 0.05.

Figure 6. 

Ureide content, allantoinase activity, and ALN1 gene expression in the seed coat at 0 (0 D) and 4 (4 D) d post-imbibition. Ureides were quantified and normalized to total soluble protein. (a) Allantoinase activity was determined by an enzymatic assay and expressed as specific activity. (b) ALN1 transcript levels were determined by qRT-PCR. (c) Relative expression levels were normalized to the geometric mean of two reference genes and then calculated using the 2^−ΔCᴛ method. Bars represent mean values ± standard error (SE) of four biological replicates. Student's t-test was performed comparing both conditions. Statistical significance levels were indicated as follows: * p < 0.05, ** p < 0.01, and *** p < 0.001.