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PD-1 immune checkpoint has been intensively investigated in pathogenesis and treatments for cancer and autoimmune diseases. Cells that express PD-1 (PD-1

Vaccines that induce cytotoxic T lymphocyte (CTL)-mediated immune responses constitute an important class of medical tools to fend off diseases like infections and malignancy. Epitope peptides, as a format of CTL vaccines, are being tested preclinically and clinically. To elicit CTL responses, epitope vaccines go through an epitope presentation pathway in dendritic cells (DCs) that has multiple bottleneck steps and hence is inefficient. Here, we report the development of a strategy to overcome one of these barriers, phagolysosomal escape in DCs. First, we furnished a previously established carrier—an immune-tolerant elastin-like polypeptide nanoparticle (iTEP NP)—with the peptides that are derived from the DNA polymerase of herpes simplex virus 1 (Pol peptides). Pol peptides were reported to facilitate phagolysosomal escape. In this study, while we found that Pol peptides promoted the CTL epitope presentation; we also discovered Pol peptides disrupted the formation of the iTEP NP. Thus, we engineered a series of new iTEPs and identified several iTEPs that could accommodate Pol peptides and maintain their NP structure at the same time. We next optimized one of these NPs so that its stability is responsive to its redox environment. This environment-responsive NP further strengthened the CTL epitope presentation and CTL responses. Lastly, we revealed how this NP and Pol peptides utilized biological cues of phagolysosomes to realize phagolysosomal escape and epitope release. In summary, we developed iTEP NP carriers with a new phagolysosomal escape function. These carriers, with their priorly incorporated functions, resolve three bottleneck issues in the CTL epitope presentation pathway: vaccine uptake, phagolysosomal escape, and epitope release.

Myotonic dystrophy type 1 (DM1) is caused by the expansion of (CTG)n in the 3′ untranslated region of the dystrophia myotonica-protein kinase (DMPK) gene, which is transcribed as (CUG)n repeats that accumulate in the nucleus. The RNA repeats specifically sequester or change the expression levels of several RNA-binding proteins, leading to aberrant splicing of many target genes. In this study, we developed artificial site-specific RNA endonucleases (ASREs) that specifically bind and cleave (CUG)n repeats RNA. We have generated one ASRE that can target the expanded RNA repeats in DM1 patient cells and specifically degrade the pathogenic DMPK messenger RNAs with minimal effect on wild-type alleles. Such ASRE treatment significantly decreased the number of nuclear foci in DM1 patient cells and can reverse the missplicing of many genes affected in DM1 patients. Taken together, the application of ASRE provides a new route of gene therapy for DM1 treatment.

Autoregulatory loops often provide precise control of the level of expression of specific genes that encode key regulatory proteins. Here we have defined a pathway by which Yra1p, a yeast mRNA export factor, controls its own expression. We show that YRA1 exon 1 sequences in cis and Yra1p in trans inhibit YRA1 pre-mRNA splicing and commit the pre-mRNA to nuclear export. Mex67p and Crm1p jointly promote YRA1 pre-mRNA export, and once in the cytoplasm, the pre-mRNA is degraded by a 5' to 3' decay mechanism that is dependent on the decapping activator Edc3p and on specific sequences in the YRA1 intron. These results illustrate how common steps in the nuclear processing, export, and degradation of a transcript can be uniquely combined to control the expression of a specific gene and suggest that Edc3p-mediated decay may have additional regulatory functions in eukaryotic cells.

Transcripts regulated by the yeast nonsense-mediated and 5' to 3' mRNA decay pathways were identified by expression profiling of wild-type, upf1Delta, nmd2Delta, upf3Delta, dcp1Delta, and xrn1Delta cells. This analysis revealed that inactivation of Upf1p, Nmd2p, or Upf3p has identical effects on global RNA accumulation; inactivation of Dcp1p or Xrn1p exhibits both common and unique effects on global RNA accumulation but causes upregulation of only a small fraction of transcripts; and the majority of transcripts upregulated in upf/nmd strains are also upregulated to similar extents in dcp1Delta and xrn1Delta strains. Our results define the core transcripts regulated by NMD, identify several novel structural classes of NMD substrates, demonstrate that nonsense-containing mRNAs are primarily degraded by the 5' to 3' decay pathway even in the absence of functional NMD, and indicate that 3' to 5' decay, not 5' to 3' decay, may be the major mRNA decay activity in yeast cells.

Cadmium (Cd) is a well-known environmental carcinogen and immunotoxin. Currently the direct cytotoxic effects of Cd on thymocytes are largely unexplored. The main objective of the present study was to investigate the apoptogenic property of Cd and the mechanisms involved, using primary cultured mouse thymocytes as a model. Cd-induced apoptosis in thymocytes was studied by TdT-mediated dUTP nick end-labeling assay and DNA gel electrophoresis. The results showed that Cd was able to cause apoptosis in mouse thymocytes in a time- and dose-dependent manner. Moreover, Cd exposure led to a rapid and sustained intracellular calcium (Ca2+) elevation, followed by caspase-3 activation and PARP cleavage, all of which preceded the characteristic DNA fragmentation. BAPTA-AM, a specific intracellular Ca2+ chelator, abolished Cd-induced Ca2+ overloading and subsequently inhibited caspase-3 activation, PARP cleavage, and apoptosis. It is believed that intracellular Ca2+ elevation may trigger caspase-3 activation either through mitochondria or through activation of Ca2+-dependent protease in Cd-treated thymocytes. Results from this study thus provide new information for a better understanding of the immunotoxic and immunomodulatory effects of Cd.