An autoantibody derived from mice with experimental systemic lupus erythematosus is directed against the essential splicing factor SF53/4--a possible role for large nuclear ribonucleoprotein particles in autoimmune disorders.
We have previously shown that nuclear transcripts of several pre-mRNAs can be released from nuclei of mammalian cells in a form of large nuclear ribonucleoprotein (InRNP) particles. These particles, which invariably sediment at the 200S region in sucrose gradient, contain all U small nuclear RNPs required for pre-mRNA splicing and a multitude of heterogeneous nuclear RNP proteins. From a panel of mAbs raised against the InRNP particles, a specific mAb (53/4) identified a nuclear protein of 88 kDa as an essential splicing factor (SF53/4). In a parallel independent study, mAbs were established in mice with experimental systemic lupus erythematosus (SLE), that had been induced by immunization with a murine mAb against a human anti-DNA mAb bearing the common 16/6 idiotype. One of the produced mAbs (2C5/3) recognized an 88 kDa RNP protein. In the present study we have used the following criteria to demonstrate that mAb 2C5/3 and mAb 53/4 recognize the same protein. First, mAb 2C5/3 inhibited splicing by direct addition. Second, the 88 kDa polypeptide that had been immunodepleted from HeLa cells nuclear extract by mAb 2C5/3 was recognized by mAb 53/4 in protein blots. Third, the HeLa nuclear extract depleted by mAb 2C5/3 was devoid of splicing activity and could not assemble into splicing complexes with exogenous pre-mRNA; however, splicing and spliceosome assembly activities were restored to such a defective extract by adding back the 88 kDa protein that had been purified by immunoaffinity binding to immobilized mAb 53/4.(ABSTRACT TRUNCATED AT 250 WORDS)
Ast
G
G
Department of Organic Chemistry, Weizmann Institute of Science, Rehovot, Israel.
Goldblatt
D
D
Waisman
A
A
Sperling
R
R
Mozes
E
E
Sperling
J
J
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001323:Autoantibodies / Q000276:immunology*; D002846:Chromatography, Affinity; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D008180:Lupus Erythematosus, Systemic / Q000276:immunology*; D051379:Mice; D008815:Mice, Inbred Strains; D012326:RNA Splicing / Q000502:physiology*; D011863:Radioimmunoassay; D012261:Ribonucleoproteins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D012261:Ribonucleoproteins