The products of the human leukocyte antigen subtypes HLA-B*2705 and HLA-B*2709 differ only in residue 116 (Asp vs. His) within the peptide binding groove but are differentially associated with the autoimmune disease ankylosing spondylitis (AS); HLA-B*2705 occurs in AS-patients, whereas HLA-B*2709 does not. The subtypes also generate differential T cell repertoires as exemplified by distinct T cell responses against the self-peptide pVIPR (RRKWRRWHL). The crystal structures described here show that pVIPR binds in an unprecedented dual conformation only to HLA-B*2705 molecules. In one binding mode, peptide pArg5 forms a salt bridge to Asp116, connected with drastically different interactions between peptide and heavy chain, contrasting with the second, conventional conformation, which is exclusively found in the case of B*2709. These subtype-dependent differences in pVIPR binding link the emergence of dissimilar T cell repertoires in individuals with HLA-B*2705 or HLA-B*2709 to the buried Asp116/His116 polymorphism and provide novel insights into peptide presentation by major histocompatibility antigens.

Objective To compare haplotype distribution in HLA–B27–positive patients with ankylosing spondylitis (AS) and healthy control subjects possessing either AS‐associated HLA–B27 alleles or the non–AS‐associated HLA–B*2709 allele. Methods DNA samples from 47 HLA–B27–positive patients with AS and 76 HLA–B27–positive healthy controls (19 positive and 57 negative for B*2709) living in different areas of Sardinia were collected and typed for HLA class I and class II alleles. The third exon of the B27 gene was analyzed for the presence of Asp116 or His116, which differentiates B*2709 from the other two B27 subtypes (B*2705 and B*2702) that are mostly found in Sardinia. The parents of 6 subjects positive for B*2709 were also typed for HLA class I and class II alleles. Statistical analysis was performed by Fisher's exact test. Results In Sardinia, the B27 alleles conferring susceptibility to AS appear to be more frequently carried by a haplotype (A2;B27;Cw2;DR16) that reaches its highest frequency in patients with AS (A2 80.8%, B27 100%, Cw2 83%, and DR16 74.5%). Conversely, the non–AS‐associated B*2709 allele is more frequently found together with other HLA alleles whose frequencies are inversely correlated with the disease (A32 or A30, Cw1, and DR12). Familial analysis of 6 subjects positive for HLA–B*2709 confirmed the existence of a “Sardinian” haplotype that is not associated with AS (A32;B*2709;Cw1;DR12). Conclusion In Sardinia, 2 distinct haplotypes harbor the non–AS‐associated HLA–B*2709 allele or the AS‐associated B27 alleles. Our findings are compatible with the hypothesis that other genes within the HLA region besides HLA–B27 may play some role in conferring susceptibility to AS.

Cytotoxic T lymphocytes (CTLs) are the most powerful weapon of the immune system to eliminate cells infected by intracellular parasites or tumors. However, very often, escape mechanisms overcome CTL immune surveillance by impairing the classical HLA class I antigen-processing pathway. Here, we describe a strategy for CTL activation based on the ability of Tat to mediate transcellular delivery of viral proteins encompassing HLA class I-restricted epitopes. In this system, the recombinant protein TAT-NpFlu containing the transduction domain of Tat of human immunodeficiency virus type 1 fused to the amino acid region 301 to 498 of the nucleoprotein of influenza A virus is proven to sensitize different human cells to lysis by HLA-B27-restricted, Flu 383-391-specific CTL lines. The fusion protein is processed very effectively, since a comparable biological effect is obtained with an amount of protein between 1 and 2 orders of magnitude lower than that of the synthetic peptide. Interestingly, while part of TAT-NpFlu undergoes fast and productive cleavage, a large amount of it remains intact for up to 24 h. Confocal microscopy shows that TAT-NpFlu accumulates in the trans-Golgi network (TGN), where it starts to be detectable 1 h after transduction. Using TAT-NpFlu mutants and hybrid constructs, we demonstrate that enrichment in the TGN occurs only when the carboxy-terminal region of NpFlu (amino acids 400 to 498) is present. These data disclose an unconventional route for presentation of epitopes restricted for HLA class I molecules.Cytotoxic T lymphocytes (CTLs) are key cells in immune defense against virus-infected or -transformed cells. However, very often, escape mechanisms overcome CTL immune surveillance by impairing the classical HLA class I antigen-processing pathway (10, 15). There is therefore a great interest in identifying novel routes for presentation on HLA class I molecules to be targeted for immunotherapy.For a long time it was believed that the strict division of HLA class I and class II processing was necessary to identify different physiological responses of the immune system to endogenous (HLA class I) or internalized (HLA class II) proteins. However, although exogenous proteins are normally excluded from presentation by HLA class I molecules, exceptions to this rule have been described in the last 10 years that indicate the possibility, in given circumstances, of engagement and presentation of exogenous antigens on HLA class I molecules either in vivo or in vitro (16,26). In this respect, the immunological meaning of exogenous and endogenous processing should be reevaluated, since several examples of CTLs primed in vivo by proteins present in extracellular fluids have been provided previously (27), and a different pathway for exogenous proteins could be a possible explanation. Moreover, this pathway could be the initiating step for CTL activation in antitumoral immunity and, eventually, parasite infections through cross-priming. Two mechanisms have been suggested to explain exogenous peptide loadin...

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