Regulation of the human tRNase ZS gene expression

AbstractThere are two types of tRNA 3′ processing endoribonucleases (tRNase Z): a short form (tRNase ZS) and a long form (tRNase ZL). Although the human genome contains both genes, little is known about the physiological role of tRNase ZS. We found that the human tRNase ZS gene expression appears to be post-transcriptionally regulated. Additionally, analyses for cis-regulatory elements for the tRNase ZS gene transcription suggested that transcription factors that bind to five different sites on the promoter work together to potentiate the transcription initiation. Furthermore, we found that tRNase ZS is predominantly present in the cytosol and hardly in the nucleus

Inhibition of HIV-1 gene expression by retroviral vector-mediated small-guide RNAs that direct specific RNA cleavage by tRNase ZL

The tRNA 3′-processing endoribonuclease (tRNase Z or 3′ tRNase; EC 3.1.26.11) is an essential enzyme that removes the 3′ trailer from pre-tRNA. The long form (tRNase ZL) can cleave a target RNA in vitro at the site directed by an appropriate small-guide RNA (sgRNA). Here, we investigated whether this sgRNA/tRNase ZL strategy could be applied to gene therapy for AIDS. We tested the ability of four sgRNA-expression plasmids to inhibit HIV-1 gene expression in COS cells, using a transient-expression assay. The three sgRNAs guide inhibition of HIV-1 gene expression in cultured COS cells. Analysis of the HIV-1 mRNA levels suggested that sgRNA directed the tRNase ZL to mediate the degradation of target RNA. The observation that sgRNA was localized primarily in nuclei suggests that tRNase ZL cleaves the HIV-1 mRNA when complexed with sgRNA in this location. We also examined the ability of two retroviral vectors expressing sgRNA to suppress HIV-1 expression in HIV-1-infected Jurkat T cells. sgRNA-SL4 suppressed HIV-1 expression almost completely in infected cells for up to 18 days. These results suggest that the sgRNA/tRNase ZL approach is effective in downregulating HIV-1 gene expression

Modulation of Gene Expression by Human Cytosolic tRNase ZL through 5′-Half-tRNA

A long form (tRNase ZL) of tRNA 3′ processing endoribonuclease (tRNase Z, or 3′ tRNase) can cleave any target RNA at any desired site under the direction of artificial small guide RNA (sgRNA) that mimics a 5′-half portion of tRNA. Based on this enzymatic property, a gene silencing technology has been developed, in which a specific mRNA level can be downregulated by introducing into cells a synthetic 5′-half-tRNA that is designed to form a pre-tRNA-like complex with a part of the mRNA. Recently 5′-half-tRNA fragments have been reported to exist stably in various types of cells, although little is know about their physiological roles. We were curious to know if endogenous 5′-half-tRNA works as sgRNA for tRNase ZL in the cells. Here we show that human cytosolic tRNase ZL modulates gene expression through 5′-half-tRNA. We found that 5′-half-tRNAGlu, which co-immunoprecipitates with tRNase ZL, exists predominantly in the cytoplasm, functions as sgRNA in vitro, and downregulates the level of a luciferase mRNA containing its target sequence in human kidney 293 cells. We also demonstrated that the PPM1F mRNA is one of the genuine targets of tRNase ZL guided by 5′-half-tRNAGlu. Furthermore, the DNA microarray data suggested that tRNase ZL is likely to be involved in the p53 signaling pathway and apoptosis

Risk factors for sternal wound infection

Background Although the utility of flaps for the treatment of sternal wound infections following median sternotomy has been reported for 30 years, there have been few reports on the risk factors for complications after reconstruction. The objective of this investigation was to identify factors related to complications after the reconstruction of sternal wound infections. Methods A retrospective analysis of 74 patients with reconstructive surgery after sternal wound infection over a 5-year period was performed. Clinical data including age, sex, body mass index (BMI), comorbidities, bacterial culture, previous cardiac surgery, wound depth, mortality rate, type of reconstructive procedure, and complication rate were collected. Results The patients' BMI ranged from 15.2 to 33.6 kg/m2 (mean, 23.1±3.74 kg/m2). Wound closure complications after reconstructive surgery were observed in 36.5% of the cases. The mortality rate was 2.7%. Diabetes mellitus significantly affected the rate of wound closure complications (P=0.041). A significant difference in the number of complications was seen between Staphylococcus aureus (S. aureus) and coagulase-negative Staphylococci (P=0.011). There was a correlation between harvesting of the internal thoracic artery and postoperative complications (P=0.048). The complication rates of the pectoralis major flap, rectus abdominis flap, omentum flap, a combination of pectoralis major flap and rectus abdominis flap, and direct closure were 23.3%, 33.3%, 100%, 37.5%, and 35.7%, respectively. Conclusions Diabetes mellitus, S. aureus, harvesting of the internal thoracic artery, and omentum flap were significant factors for complications after reconstruction. The omentum flap volume may be related to the complications associated with the omentum flap transfer in the present study

Antigen-specific CD8 T cells can eliminate antigen-bearing keratinocytes with clonogenic potential via an IFN-γ-dependent mechanism

The immune system surveys the skin for keratinocytes (KCs) infected by viruses or with acquired genetic damage. The mechanism by which T cells mediate KC elimination is however undefined. In this study we show that antigen-specific CD8 T cells can eliminate antigen-bearing KCs in vivo and inhibit their clonogenic potential in vitro, independently of the effector molecules perforin and Fas-ligand (Fas-L). In contrast, IFN-gamma receptor expression on KCs and T cells producing IFN-gamma are each necessary and sufficient for in vitro inhibition of KC clonogenic potential. Thus, antigen-specific cytotoxic T lymphocytes (CTLs) may mediate destruction of epithelium expressing non-self antigen by eliminating KCs with potential for self-renewal through an IFN-gamma-dependent mechanism

Structural basis for the binding of IRES RNAs to the head of the ribosomal 40S subunit

Some viruses exploit internal initiation for their propagation in the host cell. This type of initiation is facilitated by structured elements (internal ribosome entry site, IRES) upstream of the initiator AUG and requires only a reduced number of canonical initiation factors. An important example are IRES of the virus family Dicistroviridae that bind to the inter-subunit side of the small ribosomal 40S subunit and lead to the formation of elongation-competent 80S ribosomes without the help of any initiation factor. Here, we present a comprehensive functional and structural analysis of eukaryotic-specific ribosomal protein rpS25 in the context of this type of initiation and propose a structural model explaining the essential involvement of rpS25 for hijacking the ribosome

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target