No abstract

As hamster scrapie cannot infect mice, due to sequence differences in their PrP proteins, we find ''species barriers'' to transmission of the [URE3] prion in Saccharomyces cerevisiae among Ure2 proteins of S. cerevisiae, paradoxus, bayanus, cariocanus, and mikatae on the basis of differences among their Ure2p prion domain sequences. The rapid variation of the N-terminal Ure2p prion domains results in protection against the detrimental effects of infection by a prion, just as the PrP residue 129 Met/Val polymorphism may have arisen to protect humans from the effects of cannibalism. Just as spread of bovine spongiform encephalopathy prion variant is less impaired by species barriers than is sheep scrapie, we find that some [URE3] prion variants are infectious to another yeast species while other variants (with the identical amino acid sequence) are not. The species barrier is thus prion variant dependent as in mammals.[URE3] prion variant characteristics are maintained even on passage through the Ure2p of another species. Ure2p of Saccharomyces castelli has an N-terminal Q/N-rich ''prion domain'' but does not form prions (in S. cerevisiae) and is not infected with [URE3] from Ure2p of other Saccharomyces. This implies that conservation of its prion domain is not for the purpose of forming prions. Indeed the Ure2p prion domain has been shown to be important, though not essential, for the nitrogen catabolism regulatory role of the protein.

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ICG fluorescence angiography of intraoperative flap perfusion is feasible and correlates well with outcomes of postoperative MRI flap enhancement and flap necrosis. Additional study is needed to further refine the imaging technique and optimally characterize the clinical utility.

The major histocompatibility complex (MHC) class I molecule plays a crucial role in cytotoxic lymphocyte function. Functional class I MHC exists as a heterotrimer consisting of the MHC class I heavy chain, an antigenic peptide fragment, and beta2-microglobulin (beta2m). beta2m has been previously shown to play an important role in the folding of the MHC heavy chain without continued beta2m association with the MHC complex. Therefore, beta2m is both a structural component of the MHC complex and a chaperone-like molecule for MHC folding. In this study we provide data supporting a model in which the chaperone-like role of beta2m is dependent on initial binding to only one of the two beta2m interfaces with class 1 heavy chain. beta2-Microglobulin binding to an isolated alpha3 domain of the class I MHC heavy chain accurately models the biochemistry and thermodynamics of beta2m-driven refolding. Our results explain a 1000-fold discrepancy between beta2m binding and refolding of MHC1. The biochemical study of the individual domains of complex molecules is an important strategy for understanding their dynamic structure and multiple functions.

The major histocompatibility complex class I (MHC1) molecule plays a crucial role in cytotoxic lymphocyte function. ␤2-Microglobulin (␤2m) has been demonstrated to be both a structural component of the MHC1 complex and a chaperone-like molecule for MHC1 folding. ␤2m binding to an isolated ␣3 domain of MHC1 heavy chain at micromolar concentrations has been shown to accurately model the biochemistry and thermodynamics of ␤2m-driven MHC1 folding. These results suggested a model in which the chaperone-like role of ␤2m is dependent on initial binding to the ␣3 domain interface of MHC1 with ␤2m. Such a model predicts that a mutant ␤2m molecule with an intact MHC1 ␣3 domain interaction but a defective MHC1 ␣1␣2 domain interaction would block ␤2m-driven folding of MHC1. In this study we generated such a ␤2m mutant and demonstrated that it blocks MHC1 folding by normal ␤2m at the expected micromolar concentrations. Our data support an initial interaction of ␤2m with the MHC1 ␣3 domain in MHC1 folding. In addition, the dominant negative mutant ␤2m can block T-cell functional responses to antigenic peptide and MHC1.The major histocompatibility complex class I (MHC1) 1 molecule and antigenic peptide are recognized by CD8ϩ cytotoxic T-lymphocytes (CTL) in CTL activation and lysis of targets (1). The heavy chain of the MHC1 molecule can interact noncovalently with a number of other molecules in the formation of a CTL activating complex. These include the MHC1 light chain or ␤2m, the antigenic peptide fragment, the T-cell receptor (TCR), and the CD8 molecule (2). The specificity of the CTL response resides in the selective MHC1 binding of specific antigenic peptide fragments and in the TCR recognition of these antigenic peptides and MHC1 (3, 4). The MHC1 contact surface for TCR and peptide binding is formed by the ␣1 and ␣2 domains of the three-domain MHC1 heavy chain (2, 5-7).The MHC1 heavy chain ␣1 and ␣2 domains, as well as the immunoglobulin-like ␣3 domain, have been shown by x-ray crystallography (8) to interact with ␤2m, the nonpolymorphic component of the MHC1 complex. Mutations in the ␣1 (9, 10) or ␣3 (11) domains of the MHC1 heavy chain lead to changes in ␤2m binding. These studies demonstrate that the functional interaction of the MHC1 heavy chain with ␤2m occurs at multiple surfaces on different domains.In the absence of ␤2m, most MHC1 molecules are not expressed efficiently on the surface of cells (12, 13). Although some MHC1 molecules, such as murine H-2L d and H-2D b , are transported to the cell surface without ␤2m, they have diminished levels of expression (14,15). This decreased MHC1 expression is not simply because of an export requirement for fully assembled MHC1 complexes. Transfection of ␤2m-negative cells with ER-retained ␤2m was able to salvage MHC1 cell surface expression (16). MHC1 folded in the presence of this ER-retained ␤2m was exported to the cell surface without bound ␤2m. Thus ␤2m, which promotes protein folding through a transient interaction, fits the definition of a chaperone (17). Therefore,...

Cancer stem cells (CSCs) are cells within tumors that maintain the ability to self-renew, drive tumor growth, and contribute to therapeutic resistance and cancer recurrence. In this study, we investigate the role of Zinc finger and SCAN domain containing 4 (ZSCAN4) in human head and neck squamous cell carcinoma (HNSCC). The murine Zscan4 is involved in telomere maintenance and genomic stability of mouse embryonic stem cells. Our data indicate that the human ZSCAN4 is enriched for, marks and is co-expressed with CSC markers in HNSCC. We show that transient ZSCAN4 induction for just 2 days increases CSC frequency both in vitro and in vivo and leads to upregulation of pluripotency and CSC factors. Importantly, we define for the first time the role of ZSCAN4 in altering the epigenetic profile and regulating the chromatin state. Our data show that ZSCAN4 leads to a functional histone 3 hyperacetylation at the promoters of OCT3/4 and NANOG, leading to an upregulation of CSC factors. Consistently, ZSCAN4 depletion leads to downregulation of CSC markers, decreased ability to form tumorspheres and severely affects tumor growth. Our study suggests that ZSCAN4 plays an important role in the maintenance of the CSC phenotype, indicating it is a potential therapeutic target in HNSCC.

Objectives/Hypothesis Surgical management of nasopharyngeal tumors has evolved in the endoscopic era. Lateral exposure remains difficult especially near the petrous internal carotid artery and bony Eustachian tube (ET). Our study examines the need to sacrifice the vidian and greater palatine nerves in order to successfully perform en bloc endoscopic nasopharyngectomy. Methods Four cadaveric specimens (eight sides) were dissected bilaterally using a binarial, extended, endoscopic endonasal approach (EEA). Nasopharyngectomy was completed including an extended transptyergoid approach for resection of the cartilaginous ET at its junction with the bony ET. Dissection was attempted without sacrifice of the vidian or palatine nerves. Results Successful en bloc nasopharyngectomy combined with a nerve‐sparing transpterygoid approach was achieved in all specimens with successful preservation of the palatine and vidian nerves. The approach provided exposure of foramen lacerum, the petrous carotid, foramen spinosum, and foramen ovale as well as all segments of the cartilaginous Eustachian tube, Meckel's cave and the parapharyngeal carotid. There was no inadvertent exposure or injury of the internal carotid artery. Conclusion Endoscopic nasopharyngectomy combined with a nerve‐sparing transpterygoid approach allows for en bloc resection of the cartilaginous Eustachian tube and nasopharyngeal contents with broad skull base exposure and preservation of the internal carotid artery, vidian and palatine nerves. Level of Evidence VI Laryngoscope, 130:2343–2348, 2020