ITRAQ-based quantitative glutelin proteomic analysis reveals differentially expressed proteins in the physiological metabolism process during endosperm development and their impacts on yield and quality in autotetraploid rice

Lin Xian,Yanxi Long,Meng Yang,Zhixiong Chen,Jinwen Wu,Xiangdong Liu,Lan Wang

doi:10.1016/j.plantsci.2021.110859

Abstract

Autotetraploid rice, which is developed through chromosome set doubling using diploid rice, produces high-quality kernels that are rich in storage proteins. However, little information is available about the content of different proteins in autotetraploid rice and their proteomic analysis. The dynamic changes in four storage proteins, namely, albumin, globulin, prolamin, and glutelin, were analyzed in the endosperm of autotetraploid rice (AJNT-4x) and in that of its diploid counterpart (AJNT-2x) for comparison. The contents of the four proteins were all higher during endosperm development in AJNT-4x than in AJNT-2x, but their change and composition were almost the same in the two materials. Then, iTRAQ was employed to analyze the glutelin profiles of AJNT-4x and AJNT-2x at 10 DAF, 15 DAF, and 20 DAF. A total of 1326 proteins were identified in AJNT-4x and AJNT-2x using high-throughput LC–MS/MS. Among the 1326 identified proteins, there were 362 DEPs in AJNT-4x compared with AJNT-2x and 372 DEPs between different developmental stages in AJNT-4x. Eight important upregulated proteins were identified by qRT-PCR, including B8AM24, B8ARJ0, B8AQM6, A2ZCE6, and P37833. Among them, B8AM24 and B8ARJ0 were related to the lysine biosynthesis process. GO enrichment analysis revealed that the critical functions of DEPs exhibited little overlap between the 10, 15, and 20 DAF groups. Endosperm glutelin accumulation was regulated mainly by different DEPs during the early stage, and 15 DAF was a critical regulating point for glutelin accumulation. KEGG pathway analysis showed that ribosomal proteins were significantly higher in AJNT-4x than in AJNT-2x at 10 DAF, and protein processing, biosynthesis, and metabolism of amino acids were higher and more active in AJNT-4x at 15 DAF, while the peroxisome was richer in AJNT-4x at 20 DAF. The PPI network showed that ribosomal proteins gradually decreased with increasing endosperm development. These results provide new insights into dynamic glutelin expression differences during endosperm development in autotetraploid rice, which will aid in the development of rice cultivars with increased yield and improved grain nutritional quality.

Highlights

MethodsGrain weight and seed setting rate investigation and endosperm preparation
Autotetraploid rice, which is developed through chromosome set doubling using diploid rice, produces highquality kernels that are rich in storage proteins
Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that ribosomal proteins were significantly higher in AJNT-4x than in AJNT-2x at 10 days after flowering (DAF), and the protein processing, biosynthesis, and metabolism of amino acids were higher and more active in AJNT-4x at 15 DAF, while the peroxisome was richer in AJNT-4x at 20 DAF

Summary

Methods

Grain weight and seed setting rate investigation and endosperm preparation. Ten rows were planted in each of the test plots, and the row spacing was 18 cm×21 cm. A total of 70 rows were measured for seed setting in the middle of the test plots (Li et al 2011). The grains were labeled at the time of flowering and were collected [5, 6, 7, 8, 9, 10, 11, 12, 15, 18, 20,21], and 24 days after flowering (DAF) and stored at -80 °C in a freezer

Results

Discussion

Conclusion