M J Fritzler

When tissue sections are extracted with 0.1 N HCl, cellular nuclear proteins, including histones, are removed but nuclear DNA is retained. Histones can be reconstituted back to nuclear DNA in acid-extracted tissue sections so that the resulting nuclear substrate is composed only of DNA and histones and does not contain acidic nuclear protein antigens. The resulting DNA-histone tissue substrate can be used in the immunofluorescent method for specific detention of antibodies to histones. Sera from 23 patients with drug-induced lupus erythematosus (procainamide 19, isoniazid 2, nitrofurantoin 2) and 20 patients with idiopathic (not drug-induced) systemic lupus erythematosus (SLE) were studied. All 23 patients with drug-induced lupus erythematosus (LE) lost nuclear staining on acid-extracted sections. In contrast, only 12 of 20 with idiopathic SLE lost nuclear staining on acid-extracted tissues, and in the remaining 8, there was no significant fall in titer. In the drug-induced LE group, loss of nuclear staining was due to the absence of histones on the substrate because with histone-reconstituted sections, 22 of 23 again became positive for nuclear staining at titers equal to or at one doubling dilution below titers on unextracted tissues. In contrast, of the 12 idiopathic SLE sera which lost nuclear staining, only 5 regained nuclear staining on histone-reconstituted tissue sections. In idiopathic SLE, antinuclear antibodies are heterogeneous in specificities and may consist of antibodies to native DNA, histones, or nonhistone proteins. In contrast, antinuclear antibodies in drug-induced LE are less heterogeneous and mainly consist of antibodies to histones.

OBJECTIVE: To identify a Golgi complex autoantigen bound by Sjögren's syndrome (SS) autoantibodies. METHODS: Serum from a patient with secondary SS and anti-Golgi antibodies was used as a probe to isolate a complementary DNA (cDNA) insert from a HeLa cDNA library. RESULTS: A 3.7-kb cDNA encoding a 56-kd recombinant protein was immunoprecipitated by the human anti-Golgi serum and immune rabbit serum. Western blot analysis showed that the immune rabbit sera recognized a protein of 97 kd (golgin-97), suggesting that the isolated clone contained a partial cDNA. The 5' upstream sequence was obtained by rapid amplification of the cDNA ends. The complete cDNA contained 4,860 basepairs, encoding a protein with a calculated Mr of 88 kd. Antibodies to golgin-97 were found in 12 (20%) of 60 sera known to have anti-Golgi autoantibodies, and the majority of these sera (8 of 12, or 75%) were from patients who had secondary SS. CONCLUSION: Golgin-97 is a unique Golgi complex antigen that appears to be a target of SS autoantibodies.

The sera of patients with systemic lupus erythematosus (SLE) and drug-induced lupus (DIL) were used to study the antigenic epitopes on nuclear histones that bind antibodies in these sera. ELISA and immunoblotting techniques showed that antibodies from both patient groups bound all classes of intact histone: H1 greater than H2B greater than H2A greater than H3 greater than H4. The different classes of histone were enzymatically or chemically cleaved to produce a series of peptide fragments which were then used to map the reactive epitopes by ELISA and immunoblotting. Ten of 11 DIL sera and 11 of 12 SLE sera bound the carboxy and amino terminal peptides. Only one sera of each group bound to the central hydrophobic polypeptide. The reactivity of DIL sera with fractionated histone polypeptides was similar to that observed with SLE sera. This observation suggests that the histone epitopes reacting with DIL sera are no less restricted than those reacting with SLE.

In this study we report the identification of an antibody in the sera of some patients with autoimmune disease that reacted with a cytoplasmic antigen localized within the Golgi apparatus. The antibody reacted with all tissues investigated, which included pancreas, kidney, testis, liver, thymus, and spleen. In addition, it reacted with some human peripheral circulating lymphocytes, murine peritoneal macrophages, and a variety of tissue culture cell lines, which included HEp-2 cells (human epithelial carcinoma), baby hamster kidney cells, a canine thymus cell line, a primary kidney cell line, Ehrlich ascites cells, Wil-2 cells, and Raji cells. The antigen is located in the same region stained by the histochemical reaction for thiamine pyrophosphatase, thus indicating that the antigen is located within the Golgi apparatus. The antigen was not demonstrated by immunodiffusion of saline extracts of rabbit thymus, pancreas, or liver. The antigen in HEp-2 cells was resistant to RNase A, DNase I, micrococcal nuclease, and to extraction with 0.1 N HC1, but was sensitive to trypsin and Proteinase K. Eight patients with anti-Golgi antibodies have been identified. Six of the eight had systemic lupus erythematosus. Autoantibodies to a Golgi apparatus antigen might serve as a useful biologic marker to study the functional relationship of the Golgi apparatus to lymphocytes and macrophages.