AUTOANTIGENS AND THE NUCLEAR BODY

真核细胞的组成部分分为物理细胞和生物细胞

The components of eukaryotic cells are segregated into physical and functional compartments known as organelles. One specific domain or organelle within the nucleus has been designated the nuclear body. The term "nuclear bodies" (NBs) refers to 5-10 circumscribed structures, 0.2- 0.3 micromoles in diameter, that are distributed throughout the nucleus of resting cells. The long term objective of this study is to characterize the NB and investigate the role of this structure in human diseases. Three independent lines of investigation suggest that the NB warrants further study. l) A component of the NB (SP100) is a target of autoantibodies in 40% of patients with the autoimmune disease primary biliary cirrhosis. The relatively high frequency and specificity of antibodies directed against this organelle in patient with PBC suggest that the NB may be involved in the pathogenesis of this disease. 2) In patients with acute promyelocytic leukemia (APL), a translocation occurs between the gene encoding the retinoic acid receptor alpha (RARalpha) and a gene encoding a second component of the nuclear body (PML). The PML- RARalpha fusion protein disrupts the NB and is associated with dysregulated growth of leukemic cells. 3) During the course of herpes simplex virus l (HSV-1) infection, a viral protein (ICP0) associates with, and appears to disrupt, the NB. The viral protein activates gene transcription and is involved in reactivation of latent virus. It is expected that the proposed studies will provide insight into the pathophysiology of PBC, APL and HSV infection. In preliminary studies, serum from a patient with PBC was used to identify a cDNA encoding a novel protein, SP140, which may be a second autoantigen in the nuclear body. The amino acid sequence of SP140 is homologous to SP100. In addition, SP140 contains a nucleic acid-binding motif that is present in the two other proteins that are known to associate with the NB (PML and ICP0). The specific goals of this project include: 1) Investigate the relationship between SP140 and the nuclear body. Polyclonal antiserum directed against SP140 will be produced and used to determine the location of this protein in the cell. A cDNA encoding SP140 with an epitope tag will be transfected into eukaryotic cells and the potential co- localization of SP140 with components of the NB will be determined. 2) Characterize the interactions between SP140 and the previously identified components of the NB. SP140 will be prepared by in vitro translation and the interaction of SP140 with recombinant components of the NB will be investigated. 3) Identify additional components of the NB. Serum from patients with PBC will be used to identify cDNAs encoding new autoantigens in the NB. 4) Determine the clinical significance of autoantibodes directed against components of the NB in patients with PBC. Serum from patients with biopsy-documented PBC will be screened for antibodies directed against the NB.