Intracellular localisation of the Ro 52 kD auto-antigen in HeLa cells visualised with green fluorescent protein chimeras

Autoantibodies to the Ro/SSA and La/SSB antigens are found in patients,vith Sjogren's syndrome and systemic lupus erythematosus. The Ro/SSA autoantige n consists of a 52 kD and a 60 kD protein, complexed with one of four small RNA molecules, The La protein can associate with the complex, The Ro/SSA a utoantigens are present in all mammalian cells, but their intracellular loc ation is subject of controversy and their function remains unclear, To stud y the intracellular sorting and targeting of Ro 52 kD we have constructed e xpression plasmids encoding fusion proteins between the full-length Ro 52 k D protein as well as Ro 52kD fragments and the green fluorescent protein (G FP) from the jelly fish, Aequorea Victoria. The subcellular distribution of the GFP-Ro 52kD fusion proteins was investigated in transient expression e xperiments using transfected HeLa cells, The GFP-full-length Ro 52 kD fusio n protein was accumulated in the cytoplasm and excluded from the nucleus, W hen GFP was fused with the La protein, the fluorescence was located in the nucleus, Clones coding for Ro 52 kD fragments containing the hydrophilic ce ntral part of the Ro 52 kD protein gave the same intracellular location and type of cytoplasmic speckles as the full-length Ro 52 kD protein, In contr ast, both amino terminal and carboxy terminal fragments were uniformly dist ributed throughout the cell just like the GFP protein itself, These observa tions indicated a cytoplasmic location of the Ro 52kD protein and demonstra ted the crucial role of the hydrophilic domain in restricting the Ro 52 kD protein to this intracellular compartment.