Anti-56K autoantibodies are present in sera from patients with various autoimmune diseases, predominantly in sera from patients with rheumatoid arthritis, systemic lupus erythematosus, or Sjögren's syndrome. To clarify the molecular structure of this autoantigen, we isolated a 2.0-kilobase pair cDNA clone considered to encode the full-length 56K autoantigen. The longest open reading frame encodes a 505-amino acid polypeptide, with a predicted molecular mass of 54.4 kDa. The in vitro translated protein is recognized by all anti-56K positive patient sera tested. Antibodies affinity-purified using the bacterially expressed recombinant protein recognized the 56K autoantigen in a HeLa cell extract. cDNA sequencing revealed that the 56K cDNA shares a high degree of homology in both nucleotide (87%) and amino acid sequence (92.5%) with bovine annexin XI, indicating that the 56K cDNA encodes the human homologue of annexin XI, a member of the Ca(2+)-dependent phospholipid binding protein family. Anti-56K autoantibody exhibits both a cytoplasmic and a nuclear staining in immunofluorescence experiments. Patients' sera recognize preferentially the N-terminal region of the protein, which is specific for 56K/annexin XI and not shared by other annexins, indicating that the autoimmune response to 56K/annexin XI in these patients is specific for this annexin family member.

We have reconstructed a molecular genetic history of human annexins to chronicle their origins and dispersal throughout the genome. This involved the completion of chromosomal mapping, determination of ancestral relationships, and estimation of gene duplication dates. Fluorescence in situ hybridization localized human annexin XI (ANX11) to 10q22.3-q23.1 and annexin XIII (ANX13) to 8q24.1-q24.2. Orthologous annexins showed minor rate variation when calibrated to species separation times given by the fossil record, but paralogous subfamilies have diverged at fivefold variable rates. The rates and extents of sequence divergence were used to predict a mean separation time of 450 million years between vertebrate annexins, although their common ancestor may have emanated from invertebrate stock. Annexins XIII and VII formed a phylogenetically early clade, and annexins II and VIa were the most divergent members of two distinct clades. ANX6 may have been created by tandem duplication about 500 million years ago (Mya) and duplicated again to form ANX5 400 Mya, whereas ANX4 and ANX8 are proposed to be sequential duplication products from annexin XI. Vertebrate annexins thus proliferated via a cascade of gene duplications in higher metazoa to form at least three diverging groups of ubiquitous and structurally related genes. These can be distinguished by their dispersed genomic locations as well as their individual patterns of expression and partially differentiated functions.

>sp|P50995|ANX11_HUMAN Annexin A11 OS=Homo sapiens GN=ANXA11 PE=1 SV=1
MSYPGYPPPPGGYPPAAPGGGPWGGAAYPPPPSMPPIGLDNVATYAGQFNQDYLSGMAANMSGTFGGANMPNLYPGAPGA
GYPPVPPGGFGQPPSAQQPVPPYGMYPPPGGNPPSRMPSYPPYPGAPVPGQPMPPPGQQPPGAYPGQPPVTYPGQPPVPL
PGQQQPVPSYPGYPGSGTVTPAVPPTQFGSRGTITDAPGFDPLRDAEVLRKAMKGFGTDEQAIIDCLGSRSNKQRQQILL
SFKTAYGKDLIKDLKSELSGNFEKTILALMKTPVLFDIYEIKEAIKGVGTDEACLIEILASRSNEHIRELNRAYKAEFKK
TLEEAIRSDTSGHFQRLLISLSQGNRDESTNVDMSLAQRDAQELYAAGENRLGTDESKFNAVLCSRSRAHLVAVFNEYQR
MTGRDIEKSICREMSGDLEEGMLAVVKCLKNTPAFFAERLNKAMRGAGTKDRTLIRIMVSRSETDLLDIRSEYKRMYGKS
LYHDISGDTSGDYRKILLKICGGND