Highly polymorphic major histocompatibility complex (MHC) molecules are at the heart of adaptive immune responses, playing crucial roles in many kinds of disease and in vaccination. We report that breadth of peptide presentation and level of cell surface expression of class I molecules are inversely correlated in both chickens and humans. This relationship correlates with protective responses against infectious pathogens including Marek's disease virus leading to lethal tumours in chickens and human immunodeficiency virus infection progressing to AIDS in humans. We propose that differences in peptide binding repertoire define two groups of MHC class I molecules strategically evolved as generalists and specialists for different modes of pathogen resistance. We suggest that differences in cell surface expression level ensure the development of optimal peripheral T cell responses. The inverse relationship of peptide repertoire and expression is evidently a fundamental property of MHC molecules, with ramifications extending beyond immunology and medicine to evolutionary biology and conservation.DOI: http://dx.doi.org/10.7554/eLife.05345.001

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Research Article 2175 IntroductionDuring the cell cycle, ubiquitin-mediated proteolysis provides a swift and precise means to regulate the abundance of cell cycle regulatory proteins, including cyclins and cyclin-dependent kinase (Cdk) inhibitors. This mechanism for regulating the turnover of proteins is mediated through ubiquitin ligases, which transfer ubiquitin to target proteins, enabling their timely destruction (Hershko, 1997;Hershko and Ciechanover, 1998;Nakayama and Nakayama, 2006;Reed, 2006;Reed, 2003). E3 ubiquitin-ligases promote the specific attachment of poly-ubiquitin chains, which then triggers proteolysis by the 26S proteasome. SCF-type (Skp1-Cullin-F-box) E3 ubiquitin ligases are multi-subunit complexes that consisting of Skp1, Cullin, Rbx1 and an F-box protein (FBP) (Ang and Wade, 2005;Deshaies, 1999;Jackson and Eldridge, 2002;Nakayama and Nakayama, 2005;Vodermaier, 2004). It is the FBP, which has a crucial role in specifically recruiting the target substrate, usually directed by post-translational modification of the substrate (Cenciarelli et al., 1999;Hermand, 2006;Ho et al., 2008;Jin et al., 2004;Winston et al., 1999). In cell cycle regulation, several FBPs that regulate G1-S phase regulators have been intensively studied. These include the prototypical S-phase kinaseassociated protein 2 (Skp2, also known as Fbxl1) and the F-box and WD repeat domain containing 7 (Fbxw7) that, respectively, regulate the levels of the Cdk inhibitors cyclin E and cyclindependent kinase inhibitor 1B (CDKN1B; also known as and hereafter referred to as p27). In addition, three FBPs -Fbxo4, Another FBP that interacts with G1-S regulatory proteins is Fbox protein 7 (Fbxo7). In contrast to the destabilising effects of other FBPs, Fbxo7 acts as a specific assembly factor for cyclin DCdk6 complexes. Fbxo7 interacts directly with both Cdk6 and p27, and cooperatively increases cyclin D3 interactions with Cdk6 in vitro (Laman et al., 2005). In vivo, Fbxo7 overexpression in immortalised murine fibroblasts leads to their Cdk6-dependent transformation. These cells had increased levels of cyclin D-Cdk6 complexes and E2F activity, and formed tumours in athymic nude mice (Laman et al., 2005). Many mitogen and cytokine signalling pathways converge on cyclin D-Cdk activity, which -when overstimulated -promotes oncogenesis. Because of its Cdk6-dependent transforming activity, we propose that Fbxo7 functions as an oncogene.In U2OS and NIH3T3 cells, Fbxo7 has shown selectivity for Cdk6 over Cdk2 and Cdk4. Although the biochemical properties of Cdk4 and Cdk6 are similar, more-recent studies have indicated that differences can be discerned in their selective binding to cofactors (Sugimoto et al., 1999;Laman et al., 2005), preference for phosphorylation sites in pRb in vitro (Takaki et al., 2005), sensitivities to INK4 family members (Tourigny et al., 2002;Jones et al., 2007) and in vivo partnering with D-type cyclins (Ely et al., 2005). Also, in studies of knockout mice, tissue-specific defects are seen: Cdk4-knockout mice have impair...

Fbxo7 is an unusual F box protein that augments D-type cyclin complex formation with Cdk6, but not Cdk4 or Cdk2, and its over-expression has been demonstrated to transform immortalised fibroblasts in a Cdk6-dependent manner. Here we present new evidence in vitro and in vivo on the oncogenic potential of this regulatory protein in primary haematopoietic stem and progenitor cells (HSPCs). Increasing Fbxo7 expression in HSPCs suppressed their colony forming ability in vitro, specifically decreasing CD11b (Mac1) expression, and these effects were dependent on an intact p53 pathway. Furthermore, increased Fbxo7 levels enhanced the proliferative capacity of p53 null HSPCs when they were grown in reduced concentrations of stem cell factor. Finally, irradiated mice reconstituted with p53 null, but not wild-type, HSPCs expressing Fbxo7 showed a statistically significant increase in the incidence of T cell lymphoma in vivo. These data argue that Fbxo7 negatively regulates the proliferation and differentiation of HSPCs in a p53-dependent manner, and that in the absence of p53, Fbxo7 expression can promote T cell lymphomagenesis.

؉ T cell responses and cross reactive A20 Ig antibodies. Only vaccination with Ad-mCD40L-infected A20 cells produced a significant delay in tumor growth and long-term survival (p ‫؍‬ 0.0039). Stronger protective immunity to A20 challenge was generated by intravenous priming with A20 cells infected with Ad-mCD40L, Ad-mIL-2 or their combination followed by a boost immunization with A20 cells activated with syngeneic fibroblasts expressing CD40L. Compared to Ad-LacZ-infected A20 priming, the combination priming was most effective followed by Ad-mCD40L and Ad-mIL-2 (p ‫؍‬ 0.0027, p ‫؍‬ 0.0027, p ‫؍‬ 0.0163 respectively). Significant A20-specific CD8 ؉ T cell-mediated cytotoxicity was only demonstrated in splenocytes from these groups of vaccinated animals. By contrast, ELISPOT assay of splenocytes from all A20 prime/ boosted vaccinated groups demonstrated increases in ␥-interferon release by T cells elicited by in vitro stimulation either with A20 cells or another syngeneic 2PK-3 lymphoma, indicating the presence of cross reactive immunity. Similarly anti-A20 immunoglobulin antibodies generated after vaccination were not necessarily A20 idiotype-specific. Direct therapy of pre-established tumors was achieved with the combination of Ad-mCD40L and Ad-mIL-2 given at Days 4 and 8 at the tumor site with a significant long-term survival of 85% of tumor-bearing mice (p ‫؍‬ 0.0001). Our study strongly supports the use of Ad-CD40L and Ad-IL-2 combination therapy for the treatment of patients with B cell lymphoma.