TAPBPR: a new player in the MHC class I presentation pathway.

In order to provide specificity for T cell responses against pathogens and tumours, major histocompatibility complex (MHC) class I molecules present high-affinity peptides at the cell surface to T cells. A key player for peptide loading is the MHC class I-dedicated chaperone tapasin. Recently we discovered a second MHC class I-dedicated chaperone, the tapasin-related protein TAPBPR. Here, we review the major steps in the MHC class I pathway and the TAPBPR data. We discuss the potential function of TAPBPR in the MHC class I pathway and the involvement of this previously uncharacterised protein in human health and disease.C.H was supported by a Wellcome Trust PhD Studentship (Grant 089563) and L.H.B was funded by a Wellcome Trust Career Development Fellowship (Grant 085038).This is the author accepted manuscript. The final published version is available via Wiley at http://onlinelibrary.wiley.com/doi/10.1111/tan.12538/abstract;jsessionid=3D6AF64F5BD8C64E84634A4303842BE2.f04t01

Differential contribution of TAP and tapasin to HLA class I antigen expression

Expression of class I human leucocyte antigens (HLA) on the surface of malignant cells is critical for their recognition and destruction by cytotoxic T lymphocytes. Surface expression requires assembly and folding of HLA class I molecules in the endoplasmic reticulum with the assistance of proteins such as Transporter associated with Antigen Processing (TAP) and tapasin. Interferon-γ induces both TAP and tapasin so dissection of which protein contributes more to HLA class I expression has not been possible previously. In this study, we take advantage of a human melanoma cell line in which TAP can be induced, but tapasin cannot. Interferon-γ increases TAP protein levels dramatically but HLA class I expression at the cell surface does not increase substantially, indicating that a large increase in peptide supply is not sufficient to increase HLA class I expression. On the other hand, transfection of either allelic form of tapasin (R240 or T240) enhances HLA-B*5001 and HLA-B*5701 antigen expression considerably with only a modest increase in TAP. Together, these data indicate that in the presence of minimal TAP activity, tapasin can promote substantial HLA class I expression at the cell surface

All 4 di-leucine motifs in the first hydrophobic domain of the E5 oncoprotein of human papillomavirus type 16 are essential for surface MHC class I downregulation activity and E5 endomembrane localization

The E5 oncoprotein of human papillomavirus type 16 downregulates surface MHC Class I and interacts with the heavy chain of the MHC complex via the first hydrophobic domain, believed to form the first helical transmembrane region (TM1) of E5. TM1 contains 4 equally spaced di-leucine (LL1-LL4) motifs. Di-leucine motifs have been implicated in protein-protein interactions and as localization signals. To see if any of the 4 di-leucine motifs of TM1 are involved in MHC downregulation by E5, we mutated each LL pair into valine pairs (VV1-VV4), as mutation of leucine to valine is not expected to cause major structural alterations in E5. We found that all 4 mutations disrupted the intracellular location of E5 and abrogated its MHC I downregulating activity; however VV2 and VV4 mutants were still able to interact physically with the MHC I heavy chain (HC) in vitro, while VV1 and VV3 mutants had lost this activity. We conclude that LL1 and LL3 are necessary for the interaction with HC, but LL2 and LL4 are not. However all 4 LL motifs are responsible for the proper localization of E5 in the Golgi/ER, and the displacement of E5 from this location contributes to the abrogation of MHC I downregulation. LL1 and LL3 motifs are expected to be on one face of the TM1 helix and LL2 and LL4 on the opposite face. We propose that E5 interacts with HC via LL1 and LL3 and that all 4 di-leucine motifs act as a targeting signal

Brain transcriptome-wide screen for HIV-1 Nef protein interaction partners reveals various membrane-associated proteins.

HIV-1 Nef protein contributes essentially to the pathology of AIDS by a variety of protein-protein-interactions within the host cell. The versatile functionality of Nef is partially attributed to different conformational states and posttranslational modifications, such as myristoylation. Up to now, many interaction partners of Nef have been identified using classical yeast two-hybrid screens. Such screens rely on transcriptional activation of reporter genes in the nucleus to detect interactions. Thus, the identification of Nef interaction partners that are integral membrane proteins, membrane-associated proteins or other proteins that do not translocate into the nucleus is hampered. In the present study, a split-ubiquitin based yeast two-hybrid screen was used to identify novel membrane-localized interaction partners of Nef. More than 80% of the hereby identified interaction partners of Nef are transmembrane proteins. The identified hits are GPM6B, GPM6A, BAP31, TSPAN7, CYB5B, CD320/TCblR, VSIG4, PMEPA1, OCIAD1, ITGB1, CHN1, PH4, CLDN10, HSPA9, APR-3, PEBP1 and B3GNT, which are involved in diverse cellular processes like signaling, apoptosis, neurogenesis, cell adhesion and protein trafficking or quality control. For a subfraction of the hereby identified proteins we present data supporting their direct interaction with HIV-1 Nef. We discuss the results with respect to many phenotypes observed in HIV infected cells and patients. The identified Nef interaction partners may help to further elucidate the molecular basis of HIV-related diseases