Comparative antler proteome of sika deer from different developmental stages

Abstract

Antler is a special bone tissue that has the ability to regenerate completely periodically. It is the fastest growing bone in the animal kingdom. Antler provides a valuable research model for bone growth and mineralization. Antler grows longitudinally by endochondral ossification with their growth center located in its tip. Many scholars have carried out detailed studies on morphology and gene expression of antler tip. However, few scholars have analyzed the protein expression patterns of antler tip at different development stages. This study used label-free proteomics approach to analyze the protein expression dynamics of the antler tip in six developmental periods (15, 25, 45, 65, 100 and 130 days after the previous antler cast) and costal cartilage. In result, 2052 proteins were confidently quantified, including 1937 antler proteins and 1044 costal cartilage proteins. Moreover, 913 antler core proteins and 132 antler-special proteins were obtained. Besides, the stages special proteins and differentially expressed proteins (DEPs) in different development stages were analyzed. A total of 875 DEPs were determined by one-way AVOVA. It is found that the growth period (15, 25, 45 and 65 days) showed more up-regulated protein including several chondrogenesis-associated proteins (collagen types II, collagen types XI, HAPLN1, PAPSS1 and PAPSS2). In ossification stages, the up-regulated proteins related with lysosome (CTSD, CTSB, MMP9, CAII) indicated that the antler has higher bone remodeling activity. Given the up-regulated expression of immune-related molecules (S100A7, CATHL7, LTF, AZU1, ELANE and MPO), we speculate that the local immune system may contribute to the ossification of antler tip. In conclusion, proteomics technology was used to deeply analyze the protein expression patterns of antler at different development stages. This provides a strong support for the research on the molecular regulation mechanism of rapid growth and ossification of velvet antler.

Figure 1

Flowchart of proteomic analysis of sika deer antlers at different development stages. 3 4-year-old male sika deer were selected. The two antlers from first deer were harvested at 15 and 25 days respectively, the two antlers of the second deer were harvested at 45 and 65 days respectively, and the two antlers of the last deer were harvested at 100 and 130 days respectively. LC–MS/MS was performed three times per sample.

Figure 2

Pearson correlation analysis of sika deer antlers in 6 different development stages.

Figure 3

Overview of proteins identified for antler in different development stages and costal cartilage. (A) The upset plot diagram showing overlapping proteins identified from antlers in six different development stages. (B) The top nine terms of GO enrichment result for 913 antler core proteins. (C) The Venn diagram showing overlapping between antler core proteins and costal cartilage proteins.

Figure 4

The PPI network for antler-special proteins constructed by STRING and Cytoscape. (A) The whole network including 107 nodes and 289 edges. The disconnected proteins in the network did not be showed. (B) Sub-network of top ten hub proteins with their expanded network.

Figure 5

The GO enrichment result for antler stages-special proteins. (A–D) respectively represent the results of specific expressed proteins for the first four stages, the first five stages, 100 days and 130 days.

Figure 6

Global dynamics of proteins during antler development. The main function class was searched for GO (Biological process) and KEGG enrichment.