ABSTRACT
The Ro 60 kDa protein is an RNA binding molecule present both in the cytoplasm and in the nucleus. Cytoplasmic Ro 60 kDa is complexed to other proteins and to certain RNAs denoted hYRNAs. This RNA-protein complex is also known as the Ro/SSA antigen recognized by sera from patients with certain autoimmune disorders. Components interacting with the nuclear Ro 60 kDa protein fraction in mammalian cells have not been identified. To look for an association with previously known nuclear structures, rabbit antisera to the amino- and carboxy-terminal parts of the Ro 60 kDa protein were used in immunomorphological studies on HeLa cells. A strong speckled nuclear pattern and a weak cytoplasmic staining were detected. Double immunofluorescence staining with affinity purified anti-Ro 60 kDa antibodies and monoclonal antibodies recognizing the Sm and RNP antigens of the U snRNPs, displayed colocalization. Another U snRNP containing nuclear compartment, the coiled bodies, did not contain any Ro 60 kDa protein. Cells infected with a toga virus demonstrated redistribution of both U snRNP antigens and the Ro 60 kDa protein with retained colocalization. These results indicate a role for the nuclear fraction of the Ro 60 kDa protein in RNA processing.
Subject(s)
Autoantigens/analysis , Cell Nucleus/chemistry , RNA, Small Cytoplasmic , Ribonucleoproteins, Small Nuclear/chemistry , Ribonucleoproteins/analysis , Animals , Antibody Specificity , Autoantigens/immunology , Chlorocebus aethiops , Cornea/cytology , Cricetinae , Dogs , Fluorescent Antibody Technique , HeLa Cells/chemistry , HeLa Cells/ultrastructure , HeLa Cells/virology , Humans , Immunoblotting , Kidney Tubules, Distal/cytology , Peptide Fragments/immunology , Protein Structure, Tertiary , RNA Viruses/physiology , Rabbits , Recombinant Proteins/analysis , Recombinant Proteins/immunology , Ribonucleoproteins/immunology , Ribonucleoproteins, Small Nuclear/immunology , Vero Cells/chemistry , Vero Cells/ultrastructure , Vero Cells/virology , SS-B AntigenABSTRACT
The intranuclear distribution of a new antigen (F78) associated with U snRNPs (small nuclear RNA-protein complexes) was compared with that of the RNP and Sm protein antigens previously identified on individual snRNP particles. Human and rat cells were double stained with human autoantisera and mouse monoclonal antibodies. The binding of the human and mouse antibodies was detected with secondary antibodies conjugated with fluorescein and rhodamine, respectively. The resulting immunofluorescence patterns were compared by digital image analysis. The F78, RNP, and Sm antigens show speckled fluorescence patterns which overlap to a great extent. The F78 pattern, however, also contains two classes of structural elements not present in the RNP pattern. Furthermore, during mitosis expression of the F78 antigen is completely suppressed from early prophase to telophase, while the RNP and Sm antigens are found evenly distributed throughout the cytoplasm of the dividing cells.