Dataset Information

N-glycoproteome profiling of rice

ABSTRACT: Metabolic glycan labeling (MGL) is a technique that utilizes bioorthogonal chemical reporters to investigate protein glycosylation. While this method has been widely used for glycoproteomic profiling in animals, its application in N-glycoproteomic research in crops remains limited and requires further development. In this study, we employed N-azidoacetylgalactosamine (GalNAz), a click-compatible sugar, to conduct a large-scale analysis of the N-glycoproteome in rice. Using this approach, we successfully identified 426 N-glycosylation sites and 680 N-glycosylated proteins involved in various important biological processes such as metabolism and stress response. A subset of randomly selected proteins identified by mass spectrometry were validated and confirmed to be N-glycosylated, supporting the reliability of the MGL-based method for N-glycosylation identification in rice. Moreover, by combining mass spectrometry data with biochemical validation, we discovered that the core components of the ERAD-L machinery undergo N-glycosylation. Importantly, this N-glycan modification of the ERAD-L machinery is conserved across both plants and animals. Functional studies revealed the crucial role of N-glycosylation in proper protein degradation within the ERAD-L machinery. Overall, our research significantly expanded the database of N-glycoproteins in rice and presented a highly efficient and practical approach for large-scale identification of N-glycosylated proteins, which holds potential applications for identifying N-glycosylated proteins in other crops.

INSTRUMENT(S):

ORGANISM(S): Oryza Sativa (rice)

TISSUE(S): Plant Cell, Root, Leaf

SUBMITTER: Xilong Li

LAB HEAD: Xilong Li

PROVIDER: PXD056001 | Pride | 2025-05-07

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	21-076_LC0308_RICE-N-Site-1.raw	Raw
	21-128_LC0401_RICE-N-site-2.raw	Raw
	21-128_LC0401_RICE-N-site-3.raw	Raw
	Asn-_AspSites_1.txt	Txt
	Asn-_AspSites_2.txt	Txt

Items per page:

1 - 5 of 6

Publications

Chemical Glycoproteomic Profiling in Rice Seedlings Reveals N-glycosylation in the ERAD-L Machinery.

Lei Cong C Li Xilong X Li Wenjia W Chen Zihan Z Liu Simiao S Cheng Bo B Hu Yili Y Song Qitao Q Qiu Yahong Y Zhou Yilan Y Meng Xiangbing X Yu Hong H Zhou Wen W Chen Xing X Li Jiayang J

Molecular & cellular proteomics : MCP 20241121 2

As a ubiquitous and essential posttranslational modification occurring in both plants and animals, protein N-linked glycosylation regulates various important biological processes. Unlike the well-studied animal N-glycoproteomes, the landscape of rice N-glycoproteome remains largely unexplored. Here, by developing a chemical glycoproteomic strategy based on metabolic glycan labeling, we report a comprehensive profiling of the N-glycoproteome in rice seedlings. The rice seedlings are incubated wit ...[more]

PMID: 39577566

Similar Datasets

Project description:The specific N-glycans of glycoproteins play key roles in regulating protein localization, stability and activity. B7H3, an immune checkpoint molecule, is a highly N-glycosylated membrane protein. However, the key glycosylated asparagine (Asn) residues that mediate the function of the B7H3 protein are still unclear. Here we identify that among the four pairs of putative N-glycosylation sites, N-glycans attached to asparagine residues N91/309 and N104/322 are required for proper B7H3 protein folding in the endoplasmic reticulum (ER) and intracellular trafficking to the cell surface membrane. We demonstrate that mutations in these two pairs of N-glycosylation sites induce ER accumulation of B7H3 by blocking its ER-to-Golgi translocation and subsequently promote its degradation via the endoplasmic reticulum-associated protein degradation (ERAD) pathway. Additional evidence suggests that N-glycosylation at N91/309 and N104/322 of B7H3 is essential for its inhibition of T-cell proliferation and activation. More importantly, a monoclonal antibody, Ab-82, targeting B7H3 glycosylated at N91/309 and N104/322 is developed, which exhibits the ability to elicit cytotoxic T lymphocyte-mediated antitumor immunity via B7H3 internalization and degradation. Together, these findings provide novel insights into the functional significance of B7H3 glycosylation and offer a rationale for targeting glycosylated B7H3 as a potential strategy for immunotherapy. To compare the binding partners of WT, 8NQ, and N91/104/309/322Q and N91/104/309/322 B7H3, these B7H3 constructs were stably expressed in MDA-MB-231 cell lines. The B7H3 protein was purified using affinity capture purification, followed by immunoprecipitation and mass spectrometry to identify the B7H3 interactome. Ingenuity Pathway Analysis based on these mass spectrometry data showed that the binding to components of the endoplasmic reticulum-associated degradation (ERAD) pathway was greater for the N91/104/309/322Q and 8NQ mutants than for the N91/104/309/322 mutant and WT B7H3.