细胞自噬和人类疱疹病毒相互作用的研究进展

自噬是普遍存在于真核生物中的自我保护机制之一,在自噬相关蛋白的参与下形成了完整而复杂的吞噬和自身调节机制,使其防御过程有条不紊地进行。除维持细胞内成分更新外,自噬还可选择性识别并清除入侵的病原微生物,如人类疱疹病毒。人类疱疹病毒在全球范围内普遍存在,感染人群广泛。然而,在与宿主长期共存的过程中,人疱疹病毒已经进化出相当强的抗细胞自噬机制来破坏、阻滞宿主的这一清除作用。     

The first N-terminal transmembrane helix of each subunit of the antigenic peptide transporter TAP is essential for independent tapasin binding

The heterodimeric ABC transporter TAP translocates proteasomal degradation products from the cytosol into the lumen of the endoplasmic reticulum, where these peptides are loaded onto MHC class I molecules by a macromolecular peptide-loading complex (PLC) and subsequently shuttled to the cell surface for inspection by cytotoxic T lymphocytes. Tapasin recruits, as a central adapter protein, other components of the PLC at the unique N-terminal domains of TAP. We found that the N-terminal domains of human TAP1 and TAP2 can independently bind to tapasin, thus providing two separate loading platforms for PLC assembly. Moreover, tapasin binding is dependent on the first N-terminal transmembrane helix of TAP1 and TAP2, demonstrating that these two helices contribute independently to the recruitment of tapasin and associated factors.     

Transport and transporters in the endoplasmic reticulum

Enzyme activities localized in the luminal compartment of the endoplasmic reticulum are integrated into the cellular metabolism by transmembrane fluxes of their substrates, products and/or cofactors. Most compounds involved are bulky, polar or even charged; hence, they cannot be expected to diffuse through lipid bilayers. Accordingly, transport processes investigated so far have been found protein-mediated. The selective and often rate-limiting transport processes greatly influence the activity, kinetic features and substrate specificity of the corresponding luminal enzymes. Therefore, the phenomenological characterization of endoplasmic reticulum transport contributes largely to the understanding of the metabolic functions of this organelle. Attempts to identify the transporter proteins have only been successful in a few cases, but recent development in molecular biology promises a better progress in this field.     

ABC Transporters in Dynamic Macromolecular Assemblies

ATP-binding cassette (ABC) transporters constitute one of the largest families of integral membrane proteins, including importers, exporters, channels, receptors, and mechanotransducers, which fulfill a plethora of cellular tasks. ABC transporters are involved in nutrient uptake, hormone and xenobiotic secretion, ion and lipid homeostasis, antibiotic and multidrug resistance, and immunity, thus making them prime candidates for cellular regulation and pharmacological intervention. In recent years, numerous various structures of ABC transporters have been determined by X-ray crystallography or cryogenic electron microscopy. Structural and functional studies revealed that various auxiliary domains play key roles for the subcellular localization of ABC transporters and recruitment of regulatory factors. In this regard, the ABC transporter associated with antigen processing TAP stands out. In the endoplasmic reticulum membrane, TAP assembles the peptide-loading complex, which serves as a central checkpoint in adaptive immunity. Here, we discuss the various aspects of auxiliary domains for ABC transporter function with a particular emphasis on the structure of the peptide-loading complex, which is crucial for antigen presentation in adaptive immunity.     

The N-terminal Proline Hinge Motif Controls the Structure of Bovine Herpesvirus 1-encoded Inhibitor of the Transporter Associated with Antigen Processing Required for its Immunomodulatory Function

Due to unique features, proline residues may control protein structure and function. Here, we investigated the role of ⁵²PPQ⁵⁴ residues, indicated by the recently established experimental 3D structure of bovine herpesvirus 1-encoded UL49.5 protein as forming a characteristic proline hinge motif in its N-terminal domain. UL49.5 acts as a potent inhibitor of the transporter associated with antigen processing (TAP), which alters the antiviral immune response. Mechanisms employed by UL49.5 to affect TAP remain undetermined on a molecular level. We found that mutations in the ⁵²PPQ⁵⁴ region had a vast impact on its immunomodulatory function, increasing cell surface MHC class I expression, TAP levels, and peptide transport efficiency. This inhibitory effect was specific for UL49.5 activity towards TAP but not towards the viral glycoprotein M. To get an insight into the impact of proline hinge modifications on structure and dynamics, we performed all-atom and coarse-grained molecular dynamics studies on the native protein and PPQ mutants. The results demonstrated that the proline hinge sequence with its highly rigid conformation served as an anchor into the membrane. This anchor was responsible for the structural and dynamical behavior of the whole protein, constraining the mobility of the C-terminus, increasing the mobility of the transmembrane region, and controlling the accessibility of the C-terminal residues to the cytoplasmic environment. Those features appear crucial for TAP binding and inhibition. Our findings significantly advance the structural understanding of the UL49.5 protein and its functional regions and support the importance of proline motifs for the protein structure.