PS9-3. Differential Protein Abundance Analysis in the Longissimus Thoracis Muscle of Nellore Beef Calves in Response to Creep-Feeding.

Abstract The aim of this study was to evaluate the proteome of the Longissimus thoracis muscle of Nellore calves, comparing animals supplemented or not during the cow-calf phase. The hypothesis of the study is that early supplementation alters the protein expression pattern, influencing muscle development and metabolism, with a positive impact on productive aspects. Ninety-six male Nellore calves (60 days of age, with an average body weight of 90 kg) were used in a randomized complete block design and assigned to one of two treatments during the suckling phase (180 days): mineral salt (MS; control; ad libitum; n = 48) or corn-based protein-energy creep-feeding supplement (CF; 5 g/kg BW/day; n = 48). On day 240, Longissimus thoracis (LT) muscle samples were collected by biopsy. Samples from a subset of 24 animals (12 MS; 12 CF) were subjected to label-free LC-MS/MS shotgun mass spectrometry. Data were analyzed in the R Software using DEP, limma, and SVA packages. A total of 2,675 proteins were identified, of which 39 were classified as differentially abundant proteins (FDR < 0.1; |log2FC| > 1) between treatments. Functional enrichment analysis of the identified proteins was performed on the FLAME (V2.0) platform, using Bos taurus as the reference genome. For data integration, the STRING, WebGestalt, g:Profiler, and Enrichr tools were used. Enrichment analyses included biological processes (GO:BP) and metabolic pathways from the KEGG, Reactome, and WikiPathways databases, considering a statistical significance threshold of combined P-value < 0.05. Contrasting the control group, creep-feeding reduced PRKAR1A, ALDOC and QDPR proteins, suggesting greater sensitivity to hormonal and nutritional signals, enriching Metabolism (BTA-1430728), regulation of protein phosphorylation (GO:0001932), and regulation of phosphorus metabolism (GO:0051174) pathways. Supplementation upregulated proteins linked to mitochondrial metabolism (DLST, IDH3B, NDUFS2 and GOT2), resulting in the enrichment of pathways such as Carbon metabolism (bta01200), Biosynthesis of amino acids (bta01230), and Citric acid cycle (R-BTA-71403), indicating changes in carbon and amino acid metabolism. Cytoskeleton organization pathways were also modulated, enriching actin filament organization (GO:0007015) and supramolecular fiber organization (GO:0097435). Control group was characterized by upregulation of unconventional myosins (MYO1B, MYO1C and MYO1D), whereas creep-feeding increased actin polymerization proteins (CFL1 and CAPZA1). In conclusion, supplementation of Nellore calves during the suckling phase via creep-feeding was associated with alterations in the proteome of the Longissimus thoracis muscle, suggesting that early supplementation may be used as a nutritional strategy. These findings point toward a potential reprogramming of muscle metabolism, with increased activity of energyproducing pathways, potential modulation of cAMPPKA signaling, and cytoskeletal remodeling, suggesting metabolic and structural adaptation of skeletal muscle. Funding from the São Paulo Research Foundation (FAPESP): 2023/10712-0 and 2024/23763-4.