Biomedical subjects

Long-Yuan Li

Publications and source records attributed to Long-Yuan Li.

4 recordsLinked to original sources

PubMed2006

Tyrosine phosphorylation controls PCNA function through protein stability.

The proliferating cell nuclear antigen (PCNA) is an essential protein for DNA replication and damage repair. How its function is controlled remains an important question. Here, we show that the chromatin-bound PCNA protein is phosphorylated on Tyr 211, which is required for maintaining its function on chromatin and is dependent on the tyrosine kinase activity of EGF receptor (EGFR) in the nucleus. Phosphorylation on Tyr 211 by EGFR stabilizes chromatin-bound PCNA protein and associated functions. Consistently, increased PCNA Tyr 211 phosphorylation coincides with pronounced cell proliferation, and is better correlated with poor survival of breast cancer patients, as well as nuclear EGFR in tumours, than is the total PCNA level. These results identify a novel nuclear mechanism linking tyrosine kinase receptor function with the regulation of the PCNA sliding clamp.

Cell Nucleus↗

PubMed2006

Human cellular protein VRK2 interacts specifically with Epstein-Barr virus BHRF1, a homologue of Bcl-2, and enhances cell survival.

BHRF1, an early gene product of Epstein-Barr virus (EBV), is structurally and functionally homologous to Bcl-2, a cellular anti-apoptotic protein. BHRF1 has been shown to protect cells from apoptosis induced by numerous external stimuli. Nasopharyngeal carcinoma is an epithelial cancer associated closely with EBV infection. Specific proteins that might interact with and modulate the BHRF1 anti-apoptotic activity in normal epithelial cells are of interest. Therefore, a cDNA library derived from normal human foreskin keratinocytes was screened by the yeast two-hybrid system and a cellular gene encoding human vaccinia virus B1R kinase-related kinase 2 (VRK2) was isolated. Interaction between the cellular VRK2 and viral BHRF1 proteins was further demonstrated by glutathione S-transferase pull-down assays, confocal laser-scanning microscopy and co-immunoprecipitation. Analyses of VRK2-deletion mutants revealed that a 108 aa fragment at the C terminus was important for VRK2 to interact with BHRF1. For BHRF1, aa 1-18 and 89-142 were crucial in interacting with VRK2 and these two regions are counterparts of Bcl-2 homology domains 4 and 1. Overexpressed VRK2 alone showed a modest effect in anti-apoptosis and appeared to enhance cell survival in the presence of BHRF1. However, this enhancement was not observed when VRK2 was co-expressed with Bcl-2. The results indicate that human VRK2 interacts specifically with EBV BHRF1 and that the interaction is involved in protecting cells from apoptosis.

Cell Line↗

PubMed2005

Endosomal transport of ErbB-2: mechanism for nuclear entry of the cell surface receptor.

The cell membrane receptor ErbB-2 migrates to the nucleus. However, the mechanism of its nuclear translocation is unclear. Here, we report a novel mechanism of its nuclear localization that involves interaction with the transport receptor importin beta1, nuclear pore protein Nup358, and a host of players in endocytic internalization. Knocking down importin beta1 using small interfering RNA oligonucleotides or inactivation of small GTPase Ran by RanQ69L, a dominant-negative mutant of Ran, causes a nuclear transport defect of ErbB-2. Mutation of a putative nuclear localization signal in ErbB-2 destroys its interaction with importin beta1 and arrests nuclear translocation, while inactivation of nuclear export receptor piles up ErbB-2 within the nucleus. Additionally, blocking of internalization by a dominant-negative mutant of dynamin halts its nuclear localization. Thus, the cell membrane-embedded ErbB-2, through endocytosis using the endocytic vesicle as a vehicle, importin beta1 as a driver and Nup358 as a traffic light, migrates from the cell surface to the nucleus. This novel mechanism explains how a receptor tyrosine kinase on the cell surface can be translocated into the nucleus. This pathway may serve as a general mechanism to allow direct communication between cell surface receptors and the nucleus, and our findings thus open a new era in understanding direct trafficking between the cell membrane and nucleus.

Active Transport, Cell Nucleus↗

PubMed2004

Characterization of monoclonal antibody to the Epstein-Barr virus BHRF1 protein, a homologue of Bcl-2.

A monoclonal antibody (MAb), designated 3E8, was produced against the Epstein-Barr virus BHRF1 which is a viral homologue of the anti-apoptotic protein Bcl-2. The MAb recognized the BHRF1 protein in extracts from EBV-containing cell lines after activation and EBV-negative cell lines transfected by the BHRF1 gene. Epitope mapping by Western blot analysis revealed that the antibody bound region encompassing amino acid residues 28-33 of the BHRF1. In addition to immunoblotting, the MAb could be applied widely in detection of the BHRF1 in many assays, including immunofluorescence assay, immunohistochemistry, enzyme-linked immunosorbent assay and immunoprecipitation. Most of all, when used in immunoprecipitation experiments, the MAb 3E8 showed a better effect than the existing anti-BHRF1 MAbs since radioactive isotopes were not required to intensify signals of its target antigen. Based on its great use in a variety of immunological reactions, it is a powerful tool to elucidate the biological functions of BHRF1.

Animals↗