TIGIT/CD226 Axis Regulates Anti-Tumor Immunity.

Tumors escape immune surveillance by inducing various immunosuppressive pathways, including the activation of inhibitory receptors on tumor-infiltrating T cells. While monoclonal antibodies (mAbs) blocking programmed cell death 1 (PD-1), programmed death-ligand 1 (PD-L1), and cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) have been approved for multiple cancer indications, only a subset of patients benefit from immune checkpoint blockade therapies, highlighting the need for additional approaches. Therefore, the identification of new target molecules acting in distinct or complementary pathways in monotherapy or combination therapy with PD-1/PD-L1 blockade is gaining immense interest. T cell immunoreceptor with Ig and immunoreceptor tyrosine-based inhibitory motif (ITIM) domains (TIGIT) has received considerable attention in cancer immunotherapy. Recently, anti-TIGIT mAb (tiragolumab) has demonstrated promising clinical efficacy in non-small cell lung cancer treatment when combined with an anti-PD-L1 drug (Tecentriq), leading to phase III trial initiation. TIGIT is expressed mainly on T and natural killer cells; it functions as an inhibitory checkpoint receptor, thereby limiting adaptive and innate immunity. CD226 competes for binding with the same ligands with TIGIT but delivers a positive stimulatory signal to the immune cells. This review discusses the recent discoveries regarding the roles of TIGIT and CD226 in immune cell function and their potential application in cancer immunotherapy.

Viral hijacking of the TENT4-ZCCHC14 complex protects viral RNAs via mixed tailing.

TENT4 enzymes generate 'mixed tails' of diverse nucleotides at 3' ends of RNAs via nontemplated nucleotide addition to protect messenger RNAs from deadenylation. Here we discover extensive mixed tailing in transcripts of hepatitis B virus (HBV) and human cytomegalovirus (HCMV), generated via a similar mechanism exploiting the TENT4-ZCCHC14 complex. TAIL-seq on HBV and HCMV RNAs revealed that TENT4A and TENT4B are responsible for mixed tailing and protection of viral poly(A) tails. We find that the HBV post-transcriptional regulatory element (PRE), specifically the CNGGN-type pentaloop, is critical for TENT4-dependent regulation. HCMV uses a similar pentaloop, an interesting example of convergent evolution. This pentaloop is recognized by the sterile alpha motif domain-containing ZCCHC14 protein, which in turn recruits TENT4. Overall, our study reveals the mechanism of action of PRE, which has been widely used to enhance gene expression, and identifies the TENT4-ZCCHC14 complex as a potential target for antiviral therapeutics.

Mixed tailing by TENT4A and TENT4B shields mRNA from rapid deadenylation.

RNA tails play integral roles in the regulation of messenger RNA (mRNA) translation and decay. Guanylation of the poly(A) tail was discovered recently, yet the enzymology and function remain obscure. Here we identify TENT4A (PAPD7) and TENT4B (PAPD5) as the enzymes responsible for mRNA guanylation. Purified TENT4 proteins generate a mixed poly(A) tail with intermittent non-adenosine residues, the most common of which is guanosine. A single guanosine residue is sufficient to impede the deadenylase CCR4-NOT complex, which trims the tail and exposes guanosine at the 3' end. Consistently, depletion of TENT4A and TENT4B leads to a decrease in mRNA half-life and abundance in cells. Thus, TENT4A and TENT4B produce a mixed tail that shields mRNA from rapid deadenylation. Our study unveils the role of mixed tailing and expands the complexity of posttranscriptional gene regulation.

Terminal Uridylyltransferases Execute Programmed Clearance of Maternal Transcriptome in Vertebrate Embryos.

During the maternal-to-zygotic transition (MZT), maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However, it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here, through genome-wide profiling of RNA abundance and 3' modification, we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 (TUT4/7) are depleted in zebrafish and Xenopus, maternal mRNA clearance is significantly delayed, leading to developmental defects during gastrulation. Short-tailed mRNAs are selectively uridylated by TUT4/7, with the highly uridylated transcripts degraded faster during the MZT than those with unmodified poly(A) tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation, thereby allowing the progression of early development.

Association Between a Polymorphism (rs2071214) in Baculoviral IAP Repeat Containing 5 Gene (BIRC5) and Ischemic Stroke in Korean Population.

OBJECTIVE: To investigate whether baculoviral inhibitor of apoptosis (IAP) repeat containing 5 gene (BIRC5) polymorphisms are associated with the development and clinical phenotypes of ischemic stroke in Korea population. METHODS: We enrolled 121 ischemic stroke patients and 291 control subjects. Ischemic stroke patients were divided into subgroups according to the scores of National Institutes of Health Stroke Survey (<6 or ≥6) and Modified Barthel Index (<60 or ≥60). Single nucleotide polymorphisms (SNPs) of BIRC5 (rs3764383 and rs2071214) were selected and genotyped by direct sequencing for all subjects. Multiple logistic regression models (codominant 1 and 2, dominant, recessive, overdominant and log-additive) were used to estimate odds ratios (ORs), 95% confidence intervals (CIs), and p-values. RESULTS: In analysis of stroke susceptibility, the genotype and allele frequencies of rs3764383 exhibited no difference between the control group and the ischemic stroke group. SNP rs2071214 was associated with ischemic stroke in the codominant (p=0.003), dominant (p=0.002), overdominant (p=0.005), and log-additive (p=0.008) models, respectively. The G allele frequency of rs2071214 was significantly (p=0.009) associated with susceptibility for ischemic stroke (OR, 1.57; 95% CI, 1.12-2.21). However, in the analysis for clinical phenotype, no SNP of the BIRC5 gene was found to be associated with ischemic stroke. CONCLUSION: These results suggest that a missense SNP (rs2071214) of BIRC5 may be associated with the development of ischemic stroke in the Korean population.

Deletion of human tarbp2 reveals cellular microRNA targets and cell-cycle function of TRBP.

TRBP functions as both a Dicer cofactor and a PKR inhibitor. However, the role of TRBP in microRNA (miRNA) biogenesis is controversial and its regulation of PKR in mitosis remains unexplored. Here, we generate TRBP knockout cells and find altered Dicer-processing sites in a subset of miRNAs but no effect on Dicer stability, miRNA abundance, or Argonaute loading. By generating PACT, another Dicer interactor, and TRBP/PACT double knockout (KO) cells, we further show that TRBP and PACT do not functionally compensate for one another and that only TRBP contributes to Dicer processing. We also report that TRBP is hyperphosphorylated by JNK in M phase when PKR is activated by cellular double-stranded RNAs (dsRNAs). Hyperphosphorylation potentiates the inhibitory activity of TRBP on PKR, suppressing PKR in M-G1 transition. By generating human TRBP KO cells, our study clarifies the role of TRBP and unveils negative feedback regulation of PKR through TRBP phosphorylation.

Quantitative and multiplexed microRNA sensing in living cells based on peptide nucleic acid and nano graphene oxide (PANGO).

MicroRNA (miRNA) is an important small RNA which regulates diverse gene expression at the post-transcriptional level. miRNAs are considered as important biomarkers since abnormal expression of specific miRNAs is associated with many diseases including cancer and diabetes. Therefore, it is important to develop biosensors to quantitatively detect miRNA expression levels. Here, we develop a nanosized graphene oxide (NGO) based miRNA sensor, which allows quantitative monitoring of target miRNA expression levels in living cells. The strategy is based on tight binding of NGO with peptide nucleic acid (PNA) probes, resulting in fluorescence quenching of the dye that is conjugated to the PNA, and subsequent recovery of the fluorescence upon addition of target miRNA. PNA as a probe for miRNA sensing offers many advantages including high sequence specificity, high loading capacity on the NGO surface compared to DNA and resistance against nuclease-mediated degradation. The present miRNA sensor allowed the detection of specific target miRNAs with the detection limit as low as ~1 pM and the simultaneous monitoring of three different miRNAs in a living cell.

Cell adhesion-dependent control of microRNA decay.

Mammalian microRNAs (miRNAs) are highly stable in most cell types, and their decay mechanism remains largely unknown. Here we report that some miRNAs degrade rapidly upon the loss of cell adhesion. When cells are grown at low density or cells are detached by trypsinization or EGTA treatment, mature miR-141 is downregulated while miR-200c from a common primary transcript (pri-miR-200c∼141) remains unaffected. Blockade of transcription by Actinomycin D leads to rapid depletion of miR-141 with a half-life of <1 hr when cells are detached, indicating that the regulation occurs via RNA decay. A sequence motif (UGUCU) in the center of miR-141 is necessary for the regulation. We further find that many other miRNAs including miR-200a, miR-34a, miR-29b, miR-301a, and miR-21 are degraded upon cell splitting. Induced destruction of persistent regulatory molecules such as miRNAs may increase cellular plasticity and facilitate cellular remodeling in response to the changes in cell adhesion.