A non-coding RNA balancing act: miR-346-induced DNA damage is limited by the long non-coding RNA NORAD in prostate cancer.

BACKGROUND: miR-346 was identified as an activator of Androgen Receptor (AR) signalling that associates with DNA damage response (DDR)-linked transcripts in prostate cancer (PC). We sought to delineate the impact of miR-346 on DNA damage, and its potential as a therapeutic agent. METHODS: RNA-IP, RNA-seq, RNA-ISH, DNA fibre assays, in vivo xenograft studies and bioinformatics approaches were used alongside a novel method for amplification-free, single nucleotide-resolution genome-wide mapping of DNA breaks (INDUCE-seq). RESULTS: miR-346 induces rapid and extensive DNA damage in PC cells - the first report of microRNA-induced DNA damage. Mechanistically, this is achieved through transcriptional hyperactivation, R-loop formation and replication stress, leading to checkpoint activation and cell cycle arrest. miR-346 also interacts with genome-protective lncRNA NORAD to disrupt its interaction with PUM2, leading to PUM2 stabilisation and its increased turnover of DNA damage response (DDR) transcripts. Confirming clinical relevance, NORAD expression and activity strongly correlate with poor PC clinical outcomes and increased DDR in biopsy RNA-seq studies. In contrast, miR-346 is associated with improved PC survival. INDUCE-seq reveals that miR-346-induced DSBs occur preferentially at binding sites of the most highly-transcriptionally active transcription factors in PC cells, including c-Myc, FOXA1, HOXB13, NKX3.1, and importantly, AR, resulting in target transcript downregulation. Further, RNA-seq reveals widespread miR-346 and shNORAD dysregulation of DNA damage, replication and cell cycle processes. NORAD drives target-directed miR decay (TDMD) of miR-346 as a novel genome protection mechanism: NORAD silencing increases mature miR-346 levels by several thousand-fold, and WT but not TDMD-mutant NORAD rescues miR-346-induced DNA damage. Importantly, miR-346 sensitises PC cells to DNA-damaging drugs including PARP inhibitor and chemotherapy, and induces tumour regression as a monotherapy in vivo, indicating that targeting miR-346:NORAD balance is a valid therapeutic strategy. CONCLUSIONS: A balancing act between miR-346 and NORAD regulates DNA damage and repair in PC. miR-346 may be particularly effective as a therapeutic in the context of decreased NORAD observed in advanced PC, and in transcriptionally-hyperactive cancer cells.

A potential role for the silent information regulator 2 homologue 1 (SIRT1) in periapical periodontitis.

AIM: To investigate the role played by silent information regulator 2 homologue 1 (SIRT1) during angiogenesis of periapical periodontitis. METHODOLOGY: Periapical granulomas were subjected to dual-colour immunofluorescence imaging and real-time polymerase chain reactions assaying the expression levels of SIRT1, vascular endothelial growth factor (VEGF) and VE-cadherin. The association between Ki-67 and SIRT1 expression was also examined. Human umbilical vein endothelial cells (HUVECs) were treated with a combination of lipopolysaccharide and resveratrol (a SIRT1 activator) or sirtinol (a SIRT1 inhibitor); and the levels of mRNAs encoding SIRT1, VEGF and VE-cadherin were determined. HUVEC tube formation was assayed in the presence of resveratrol or sirtinol. The Mann-Whitney U-test or the Tukey-Kramer test was used for statistical analysis. RESULTS: Ki-67-expressing cells, including endothelial cells, lay adjacent to SIRT1-expressing cells in periapical granulomas. In addition, SIRT1-expressing cells were detected adjacent to VEGF-expressing cells and VEGF- or VE-cadherin-expressing endothelial cells. SIRT1, VEGF and VE-cadherin mRNA expression levels in periapical granulomas were significantly higher (P = 0.0054, 0.0090 and 0.0090, respectively) than those in healthy gingival tissues. HUVECs treated with resveratrol exhibited significantly higher expression of mRNAs encoding SIRT1, VEGF and VE-cadherin (P = 0.0019, 0.00005 and 0.0045, respectively) compared with controls, but sirtinol inhibited such expression. Resveratrol caused HUVECs to form tube-like structures, whilst sirtinol inhibited this process. CONCLUSIONS: These findings suggest that SIRT1 may stimulate angiogenesis in periapical granulomas by triggering the proliferation of endothelial cells and inducing VEGF and VE-cadherin expression.

Change in oral health status associated with menopause in Japanese dental hygienists.

OBJECTIVES: Oral symptoms such as xerostomia and burning mouth syndrome have been recognized to increase associated with menopause. The purpose of this study was to clarify the changes in oral health as well as systemic health due to menopause and their relations with hormonal change and mental status. METHODS: Ninety-seven female dental hygienists aged 40-59 years were assigned to premenopausal, menopausal and post-menopausal groups based on self-reported menstrual condition. Subjective health statuses were evaluated by questionnaire, and objective holistic and oral statuses were evaluated by measuring serum 17ß-estradiol (E2), salivary flow rate, α-amylase and secretory IgA (SIgA) and taste sensitivity. RESULTS: A significant difference among the three groups was observed in the self-rating questionnaire of depression (SRQ-D) score and serum E2 level as well as unstimulated salivary flow rate, whereas no significant difference was observed in Simplified menopausal index, Short-Form 36-Item Health Survey, General Oral Health Assessment Index, salivary α-amylase activity, salivary SIgA concentration and taste threshold. Serum E2 levels positively correlated with unstimulated salivary flow rates and negatively correlated with SRQ-D scores and α-amylase activities. CONCLUSIONS: The results demonstrated a negative correlation between E2 levels and SRQ-D scores as well as salivary α-amylase activities, suggesting an influence of E2 on mental condition. Furthermore, E2 decrease may result in reduction of salivary flow which in turn causes various problems of oral health. Since the participants were graduates from several dental hygienist schools and working at various places, these results can be generalized to Japanese dental hygienists to some extent.

Randomized clinical trial comparing two vessel-sealing devices with crush clamping during liver transection.

BACKGROUND: Previous RCTs have failed to demonstrate the usefulness of combining energy devices with the conventional clamp crushing method to reduce blood loss during liver transection. Here, the combination of an ultrasonically activated device (UAD) and a bipolar vessel-sealing device (BVSD) with crush clamping was investigated. METHODS: Patients scheduled to undergo hepatectomy at the University of Tokyo Hospital or Nihon University Itabashi Hospital were eligible for this parallel-group, single-blinded randomized study. Patients were assigned to a control group (no energy device used), an UAD group or a BVSD group. The primary endpoint was the volume of blood loss during liver transection. Outcomes of the control group and the combined energy device groups (UAD plus BVSD) were first compared. Pairwise comparisons among the three groups were made for outcomes for which the combined energy device group was superior to the control group. RESULTS: A total of 380 patients were enrolled between July 2012 and May 2014; 116 patients in the control group, 122 in the UAD group and 123 in the BVSD group were included in the final analysis. Median blood loss during liver transection was lower in the combined energy device group (245 patients) than in the control group (116 patients): median 190 (range 0-3575) versus 230 (range 3-1570) ml (P = 0·048). Pairwise comparison revealed that blood loss was lower in the BVSD group than in the control group (P = 0·043). CONCLUSION: The use of energy devices combined with crush clamping reduced blood loss during liver transection. Registration number: C000008372 (www.umin.ac.jp/ctr/index.htm).

The role of PSMB9 upregulated by interferon signature in the pathophysiology of cutaneous lesions of dermatomyositis and systemic lupus erythematosus.

BACKGROUND: Dermatomyositis (DM) and systemic lupus erythematosus (SLE) have common skin features, including dermal mucin deposition and interferon signature, although their roles are unknown. OBJECTIVES: To identify common or specific molecular changes in DM and SLE skin. METHODS: Proteomic analysis was performed using DM and healthy skin. Glycosaminoglycans were analysed by high-performance liquid chromatography. RESULTS: The expression of 60 proteins was upregulated or downregulated in DM skin compared with healthy skin in the proteomic analysis. Among those proteins, PSMB9, an immunoproteasome subunit, was upregulated in the epidermis of DM and SLE, but not in other skin diseases. Furthermore, versican V1, a core protein for glycosaminoglycans, was upregulated, while type I collagen was downregulated in the dermis of DM and SLE skin. Interferon stimulated PSMB9 expression in cultured keratinocytes and reduced collagen expression in dermal fibroblasts, but did not affect versican expression. The PSMB9 knock-down in keratinocytes led to significant suppression of transforming growth factor (TGF)-ß2 and TGF-ß3, inducers of versican synthesis. TGF-ß3 expression was upregulated in both DM and SLE, while TGF-ß2 expression was increased only in the DM epidermis. ΔDiHS-diS1, a component of heparan sulfate, was significantly increased only in DM. TGF-ß2 expression significantly increased the ΔDiHS-diS1 expression in dermal fibroblasts in vitro. CONCLUSIONS: The interferon signature in DM and SLE skin reduces collagen in dermal fibroblasts, whereas overexpression of PSMB9 induced by interferon stimulates versican inducers in epidermal keratinocytes. In addition, the TGF-ß2-ΔDiHS-diS1 pathway may be responsible for the specific molecular change in DM skin.

Revision of the Massarineae (Pleosporales, Dothideomycetes).

We here taxonomically revise the suborder Massarineae (Pleosporales, Dothideomycetes, Ascomycota). Sequences of SSU and LSU nrDNA and the translation elongation factor 1-alpha gene (tef1) are newly obtained from 106 Massarineae taxa that are phylogenetically analysed along with published sequences of 131 taxa in this suborder retrieved from GenBank. We recognise 12 families and five unknown lineages in the Massarineae. Among the nine families previously known, the monophyletic status of the Dictyosporiaceae, Didymosphaeriaceae, Latoruaceae, Macrodiplodiopsidaceae, Massarinaceae, Morosphaeriaceae, and Trematosphaeriaceae was strongly supported with bootstrap support values above 96 %, while the clades of the Bambusicolaceae and the Lentitheciaceae are moderately supported. Two new families, Parabambusicolaceae and Sulcatisporaceae, are proposed. The Parabambusicolaceae is erected to accommodate Aquastroma and Parabambusicola genera nova, as well as two unnamed Monodictys species. The Parabambusicolaceae is characterised by depressed globose to hemispherical ascomata with or without surrounding stromatic tissue, and multi-septate, clavate to fusiform, hyaline ascospores. The Sulcatisporaceae is established for Magnicamarosporium and Sulcatispora genera nova and Neobambusicola. The Sulcatisporaceae is characterised by subglobose ascomata with a short ostiolar neck, trabeculate pseudoparaphyses, clavate asci, broadly fusiform ascospores, and ellipsoid to subglobose conidia with or without striate ornamentation. The genus Periconia and its relatives are segregated from the Massarinaceae and placed in a resurrected family, the Periconiaceae. We have summarised the morphological and ecological features, and clarified the accepted members of each family. Ten new genera, 22 new species, and seven new combinations are described and illustrated. The complete ITS sequences of nrDNA are also provided for all new taxa for use as barcode markers.

The comparative neuroanatomy and neurochemistry of zebrafish CNS systems of relevance to human neuropsychiatric diseases.

Modulatory neurotransmitters which signal through G protein-coupled receptors control brain functions which deteriorate in degenerative brain diseases. During the past decade many of these systems have been mapped in the zebrafish brain. The main architecture of the systems in zebrafish brain resembles that of the mammals, despite differences in the development of the telencephalon and mesodiencephalon. Modulatory neurotransmitters systems which degenerate in human diseases include dopamine, noradrenaline, serotonin, histamine, acetylcholine and orexin/hypocretin. Although the number of G protein-coupled receptors in zebrafish is clearly larger than in mammals, many receptors have similar expression patterns, binding and signaling properties as in mammals. Distinct differences between mammals and zebrafish include duplication of the tyrosine hydroxylase gene in zebrafish, and presence of one instead of two monoamine oxidase genes. Zebrafish are sensitive to neurotoxins including MPTP, and exposure to this neurotoxin induces a decline in dopamine content and number of detectable tyrosine hydroxylase immunoreactive neurons in distinct nuclei. Sensitivity to important neurotoxins, many available genetic methods, rapid development and large-scale quantitative behavioral methods in addition to advanced quantitative anatomical methods render zebrafish an optimal organism for studies on disease mechanisms.

Relative importance of the area and intensity of the orange spots of male guppies Poecilia reticulata as mating traits preferred by females.

Digitally modified videos of male guppies Poecilia reticulata were used to examine the relative importance of the area and intensity of the orange spots as mating traits preferred by females. The females prioritized the area of the orange spots over intensity for their mate preference.

Molecular characterization and histochemical demonstration of salmon olfactory marker protein in the olfactory epithelium of lacustrine sockeye salmon (Oncorhynchus nerka).

Despite the importance of olfactory receptor neurons (ORNs) for homing migration, the expression of olfactory marker protein (OMP) is not well understood in ORNs of Pacific salmon (genus Oncorhynchus). In this study, salmon OMP was characterized in the olfactory epithelia of lacustrine sockeye salmon (O. nerka) by molecular biological and histochemical techniques. Two cDNAs encoding salmon OMP were isolated and sequenced. These cDNAs both contained a coding region encoding 173 amino acid residues, and the molecular mass of the two proteins was calculated to be 19,581.17 and 19,387.11Da, respectively. Both amino acid sequences showed marked homology (90%). The protein and nucleotide sequencing demonstrates the existence of high-level homology between salmon OMPs and those of other teleosts. By in situ hybridization using a digoxigenin-labeled salmon OMP cRNA probe, signals for salmon OMP mRNA were observed preferentially in the perinuclear regions of the ORNs. By immunohistochemistry using a specific antibody to salmon OMP, OMP-immunoreactivities were noted in the cytosol of those neurons. The present study is the first to describe cDNA cloning of OMP in salmon olfactory epithelium, and indicate that OMP is a useful molecular marker for the detection of the ORNs in Pacific salmon.

ANNUAL EFFECTIVE DOSE ASSESSMENT DUE TO RADON AND THORON PROGENIES IN DWELLINGS OF KILIMAMBOGO, KENYA.

Human beings are continuously exposed to ionising radiation originating from natural or artificial sources. Uranium-238 and Thorium-232 found in building materials are important sources of radon and thoron in the indoor environment. The concentration levels of radon, thoron and thoron progeny were measured in mud-walled, metallic or iron sheet-walled and stone-walled modern houses in Kilimambogo region, Kenya for 3 months. Radon and thoron concentration levels were measured using passive radon-thoron discriminative monitors (RADUET), while thoron progeny concentrations as the equilibrium equivalent thoron concentration (EETC) were measured using thoron progeny monitors. The mean radon concentration levels in mud, metallic and stone-walled dwellings were 67 ± 11, 60 ± 10 and 75 ± 10 Bq m-3, respectively. The mean thoron concentration levels in the corresponding dwellings were 195 ± 36, 71 ± 24 and 161 ± 31 Bq m-3, respectively, while EETCs were 12 ± 2, 3 ± 1 and 7 ± 1 Bq m-3, respectively. The annual effective doses for radon were 1.3 ± 0.2, 1.1 ± 0.1 and 1.4 ± 0.2 mSv y-1 in mud, metallic and stone-walled houses while those from thoron estimated from EETC were 2.4 ± 0.4, 0.5 ± 0.1 and 1.5 ± 0.2 mSv y-1 in the corresponding houses, respectively.

CHANGES OF ABSORBED DOSE RATE IN AIR BY CAR-BORNE SURVEY IN NAMIE TOWN, FUKUSHIMA PREFECTURE AFTER THE FUKUSHIMA DAIICHI NUCLEAR POWER PLANT ACCIDENT.

The latest car-borne survey was carried out by Hirosaki University in order to grasp the local distribution of the absorbed dose rate in air after the evacuation order was lifted on Namie Town in 2017. The car-borne survey of absorbed dose rate in air was carried out on most of the roads which were accessible by car in Namie Town using a 3-in × 3-in NaI(Tl) scintillation spectrometer. The range of the absorbed dose rate in air was calculated to be 0.041-11 µGy h-1. The distribution maps of the absorbed dose rate in air were drawn based on the data obtained during the surveys in 2011, 2015 and 2017. The comparison of these absorbed dose rates in air suggests that the elevated absorbed dose rate in air in Namie Town caused by the FDNPP accident may be decreasing faster than natural decline which includes weathering effect and physical decay due to the artificial decontamination.

IDENTIFICATION OF NOVEL BIOMARKERS FOR LUNG CANCER RISK IN HIGH LEVELS OF RADON BY PROTEOMICS: A PILOT STUDY.

Radon is the second most important risk factor for lung cancer after tobacco smoking. In Chiang Mai, Thailand, the values of indoor radon activity concentrations are considerably higher than global average values and it is a highest level among East Asian countries. The aim of our study is to identify novel biomarkers for lung cancer risk in high radon areas using a proteomic approach. In our transitional study, a total of 81 participants of non-smokers were examined, consist of 25 lung cancer patients (LC), 16 healthy controls from low levels of natural radiation areas (LLNRA) and 40 healthy controls from high levels of natural radiation areas (HLNRA). The results showed that a total of 799 differentially expressed proteins were identified. Among these, a total of 25 proteins were observed in both LC and HLNRA, but not in LINRA. Owing to the results obtained from this study, we also point out the research direction regarding the validation of some new candidate protein as a biomarker to screen population with high risk for lung cancer in the area with high levels of radon.

Effects of colloidal iron overload on renal and hepatic siderosis and the femur in male rats.

Genetic hemochromatosis is an iron overload disorder, and osteopenic and osteoporotic. Femoral neck bone mineral density (BMD) appears to fall with rising hepatic iron concentrations. A critical role for iron in mediating tissue injury is played via hydroxyl radical formation in nephrotoxicity. We investigated the effects of a colloidal iron overload on renal function, organ siderosis, and femoral bone in male rats. Iron overload reduced body growth, and increased the weights of the liver and spleen. Marked deposition of iron was noted in liver and kidney. Activities of lactate dehydrogenase and alkaline phosphatase were decreased, and the concentrations of blood urea nitrogen and creatinine were increased with the reduction in plasma calcium and inorganic phosphorus levels, i.e. functions of the liver and kidney might be affected by reactive oxygen species such as the superoxide radical, H2O2, and the hydroxyl radical produced by overloaded iron. Damage to the proximal tubular epithelial cells of the kidney and a loss of connectivity of cancellous bone in the epiphysis and of trabecular bone in the metaphysis of the distal femur were observed in iron-overloaded rats with a reduction of femoral bone mineral density, i.e. reabsorption of calcium from the proximal tubular epithelial cells of the kidney might be affected and urinary discharge of calcium might be elevated. It was suggested that iron overload gave rise to osteoporosis combined with renal dysfunction and liver iron overload syndrome.

Painless self-monitoring of blood glucose at finger sites.

INTRODUCTION: Skin punctures at alternative sites can reduce the pain associated with self-monitoring of blood glucose (SMBG). However, delays in detection of blood glucose at alternative sites during rapid systemic blood glucose change have been reported. Accordingly, we developed a novel method of finger pricking and tested it to see if it would both reduce or eliminate pain and accurately reflect systemic glucose levels, even during rapid changes. METHODS: Each of 35 healthy volunteers (10 females and 25 males; mean age 36.6 years, range 18-82 years) received 5 serial punctures from a lancet device on randomly selected fingers. The puncture target was the dorsal side of the finger between the nail and the distal finger joint. We used a lancet device designed for accurate punctures of 0.2, 0.3, 0.4, 0.5 and 0.6 mm depths. Immediately afterward, as a control, a conventional fingertip puncture was done on the front of a 6th finger. After each puncture, subjects were asked whether or not they felt pain. RESULTS: Following punctures at depths of 0.2 mm and 0.3 mm, respectively, 31 of 35 subjects (89%) and 33 of 35 subjects (94%) felt no pain. Following conventional punctures, all 35 subjects (100%) felt pain. Blood sample volumes> or =2microL were obtained in all cases except for two punctures at 0.2 mm depth. Blood glucose levels did not differ with differing puncture sites, (conventional fingertip sites vs. alternative sites used in this study). CONCLUSIONS: Our findings demonstrate that 0.2 and 0.3 mm punctures at our alternative finger site - the dorsal side of the finger between the nail and the distal finger joint - can provide blood samples sufficient for SMBG, substantially redu-ces the proportion of subjects who experience pain, and accurately reflects systemic glucose levels.

ABO blood type incompatible kidney transplantation without splenectomy prepared with plasma exchange and rituximab.

We have designed a protocol for ABO-incompatible kidney transplantations based on preoperative plasmapheresis with a tacrolimus/mycophenolate mofetil/methylprednisolone/basiliximab protocol using low-dose rituximab (200 mg/body) instead of splenectomy to prevent antibody-mediated acute rejection. Eight patients successfully received transplants with this protocol. The titers of anti-A and -B antibodies as well as the number of CD20(+) cells were readily maintained at a low level posttransplantation. There were no side effects. All patients have renal transplant function with a follow-up of 1-34 months.

Autologous blood transfusion for kidney transplant recipients.

Autologous blood transfusion (ABT) is rarely employed in patients with end-stage renal disease (ESRD); these patients are usually anemic. Since 1998, we have attempted ABT for ESRD patients undergoing living-related kidney transplantation. Among 20 patients enrolled in this study the preoperative hemoglobin and hematocrit levels were 10.0 +/- 1.2 mg/dL (range, 8.1-11.7) and 30.0 +/- 3.7% (range, 24.7-34.3), respectively. Blood volume collected on each occasion was 235.7 +/- 57.7 mL (range, 200-400), and the number of blood collections was 2.45 +/- 0.9 (range, 1-4). Total collected volume was 567.5 +/- 157.5 mL (range, 400-800). Symptomatic hypotension was seen in two patients, but vital signs recovered spontaneously. No other problems related to blood collection were observed. Allogeneic transfusion was need in only one patient (5%). ABT was safe and efficacious in ESRD patients scheduled for living-related kidney transplantation.

SOLVING THE INTERIOR PROBLEM OF COMPUTED TOMOGRAPHY USING A PRIORI KNOWLEDGE.

The case of incomplete tomographic data for a compactly supported attenuation function is studied. When the attenuation function is a priori known in a subregion, we show that a reduced set of measurements is enough to uniquely determine the attenuation function over all the space. Furthermore, we found stability estimates showing that reconstruction can be stable near the region where the attenuation is known. These estimates also suggest that reconstruction stability collapses quickly when approaching the set of points that are viewed under less than 180 degrees. This paper may be seen as a continuation of the work "Truncated Hilbert transform and Image reconstruction from limited tomographic data" that was published in Inverse Problems in 2006. This continuation tackles new cases of incomplete data that could be of interest in applications of computed tomography.

Page Kidney Following a Nonepisode Protocol Renal Allograft Biopsy: A Case Report.

Page kidney refers to a clinical condition that is characterized by the acute onset of hypertension and renal dysfunction owing to external compression of the kidney by a hematoma, tumor, lymphocele, or urinoma. We report a case in which Page kidney occurred after a nonepisode protocol renal allograft biopsy. A 31-year-old man with end-stage renal disease received a living related kidney transplant from his father. One year later, a nonepisode protocol renal allograft biopsy was performed. A day later, the patient's serum creatinine level increased to 4.23 mg/dL, and a subcapsular renal hematoma was detected using ultrasonography and computed tomography. Page kidney was diagnosed, and immediate surgical removal of the hematoma was performed. Nine days after the operation, the patient's serum creatinine level had improved to 1.89 mg/dL. Page kidney is a serious but treatable complication of renal allograft biopsies, and clinicians should pay attention to such complications, even in the setting of nonepisode protocol renal allograft biopsies.

Immunocytochemical analyses and targeted gene disruption of GTPBP1.

We previously identified a gene encoding a putative GTPase, GTPBP1, which is structurally related to elongation factor 1alpha, a key component of protein biosynthesis machinery. The primary structure of GTPBP1 is highly conserved between human and mouse (97% identical at the amino acid level). Expression of this gene is enhanced by gamma interferon in a monocytic cell line, THP-1. Although counterparts of this molecule in Caenorhabditis elegans and Ascaris suum have also been identified, the function of this molecule remains to be clarified. In the present study, our immunohistochemical analyses on mouse tissues revealed that GTPBP1 is expressed in some neurons and smooth muscle cells of various organs as well as macrophages. Immunofluorescence analyses revealed that GTPBP1 is localized exclusively in cytoplasm and shows a diffuse granular network forming a gradient from the nucleus to the periphery of the cells in smooth muscle cell lines and macrophages. To investigate the physiological role of GTPBP1, we used targeted gene disruption in embryonic stem cells to generate GTPBP1-deficient mice. The mutant mice were born at the expected Mendelian frequency, developed normally, and were fertile. No manifest anatomical or behavioral abnormality was observed in the mutant mice. Functions of macrophages, including chemotaxis, phagocytosis, and nitric oxide production, in mutant mice were equivalent to those seen in wild-type mice. No significant difference was observed in the immune response to protein antigen between mutant mice and wild-type mice, suggesting normal function of antigen-presenting cells of the mutant mice. The absence of an eminent phenotype in GTPBP1-deficient mice may be due to functional compensation by GTPBP2, a molecule we recently identified which is similar to GTPBP1 in structure and tissue distribution.

HSP47: a tissue-specific, transformation-sensitive, collagen-binding heat shock protein of chicken embryo fibroblasts.

We report the isolation and characterization of a cDNA clone encoding HSP47, a transformation-sensitive heat shock protein that binds to collagen. A cDNA library was prepared from total RNA isolated from heat-shocked chicken embryo fibroblasts and screened by using oligonucleotide mixtures prepared on the basis of the N-terminal amino acid sequence of biochemically purified HSP47. The cDNA insert contained 3,278 bp, which encoded a 15-amino-acid signal peptide and a mature protein coding region consisting of 390 amino acid residues; it also included part of the 5' noncoding region and a long 3' noncoding region. The deduced amino acid sequence revealed an RDEL sequence at the C terminus, which is a variant of the KDEL retention signal for retention of proteins in the endoplasmic reticulum. Northern (RNA) blot analyses and nuclear run-on assays established that the induction of HSP47 by heat shock and its suppression after transformation of chicken embryo fibroblasts by Rous sarcoma virus are regulated at the transcriptional level. A homology search revealed that this protein belongs to the serpin family, the superfamily of plasma serine protease inhibitors. Although structurally homologous to the serpins, HSP47 lacks the active site thought to be essential for the inhibition of proteases and does not appear to bind to intracellular proteases. HSP47 is the first heat shock protein found to be a member of the serpin superfamily. Conversely, it is the first serpin family member that is not secreted from cells, which could be explained by acquisition of the RDEL retention signal during evolution.