Synthetic Oligonucleotide Probes Research Articles

Vsp antigens and vsp-related DNA sequences in field isolates of Mycoplasma bovis.

The expression of the 1E5 epitope which is common to the three characterized variable lipoproteins VspA, VspB and VspC of Mycoplasma bovis type strain PG45 and the presence of vsp gene DNA sequences were assessed in field isolates randomly collected from cattle showing clinical manifestations due to M. bovis infection. Among 250 isolates tested, only four failed to react with mAb 1E5. Southern blot analysis of these four isolates and of 20 isolates expressing the 1E5 epitope were performed using synthetic oligonucleotide probes corresponding to a sequence located in the Vsp signal peptide coding region common to all known Vsp products or to selected regions of previously characterized vsp genes, vspA, vspE and vspF. The results demonstrate the presence of multiple vsp-related DNA sequences in all M. bovis field isolates tested and indicate that the vsp repertoire varies in size and composition among isolates.

Vsp antigens and vsp-related DNA sequences in field isolates of Mycoplasma bovis

The expression of the 1E5 epitope which is common to the three characterized variable lipoproteins VspA, VspB and VspC of Mycoplasma bovis type strain PG45 and the presence of vsp gene DNA sequences were assessed in field isolates randomly collected from cattle showing clinical manifestations due to M. bovis infection. Among 250 isolates tested, only four failed to react with mAb 1E5. Southern blot analysis of these four isolates and of 20 isolates expressing the 1E5 epitope were performed using synthetic oligonucleotide probes corresponding to a sequence located in the Vsp signal peptide coding region common to all known Vsp products or to selected regions of previously characterized vsp genes, vspA, vspE and vspF. The results demonstrate the presence of multiple vsp-related DNA sequences in all M. bovis field isolates tested and indicate that the vsp repertoire varies in size and composition among isolates.

Molecular cloning of Soli E2: an elastase-like serine proteinase from the imported red fire ant ( Solenopsis invicta)

The 4th instar larva of the imported red fire ant, Solenopsis invicta, possesses several serine proteinases, one of which, referred to as Soli E2, has been found to contain elastase-like activity. A cDNA library was constructed in λgt10 using Poly(A) +RNA from these larvae and a synthetic degenerate oligonucleotide probe corresponding to a sequence encoding the N-terminal of Soli E2 was utilized to screen the library. Two clones were obtained covering the entire coding region of Soli E2, and the predicted structure of this proteinase indicated that it was synthesized as a preproenzyme of 249 amino acids, which was cleaved to give the mature form of 226 residues. In addition, both clones showed identity to the N-terminal amino acid sequence obtained from Edman degradation of the mature purified proteinase. Significantly, Northern blot analysis of the different developmental stages revealed that Soli E2 was selectively expressed in the 4th instar larvae.

WITHIN-COLONY RELATEDNESS IN A TERMITE SPECIES: GENETIC ROADS TO EUSOCIALITY?

Abstract Hamilton's theory predicts that relatedness asymmetries, with higher relatedness between alloparents and brood than between parents and brood, favour the evolution of eusociality. The haplodiploid reproductive system of the social Hymenoptera does indeed produce relatedness asymmetries, but the diplodiploid system of the eusocial Isoptera does not automatically do so. Three mechanisms that might favour relatedness asymmetries, and therefore eusociality, in termites have been extensively debated: First, substantial inbreeding generates the background for effective kin-selection. Second, inbreeding-outbreeding cycles within and between colonies cause a higher relatedness between individuals of the same generation than between them and their potential offspring. This would be analogous to the haplodiploid system. Third, translocation complexes of sex-linked chromosomes may generate higher relatedness within sexes than between sexes, again analogous to the haplodiploid system. We tested these three hypotheses for the African termite Schedorhinotermes lamanianus (Isoptera, Rhinotermitidae) using estimates of within-colony relatedness derived by multilocus DNA fingerprinting with a synthetic oligonucleotide probe. We found little support for any of the three hypotheses. We observed inbreeding to occur only during one or a few generations within colonies, which is unlikely to be an operational basis for ongoing kin-selection. Overall, we conclude that ecological factors and constraints must be considered a major selective force.

Identification of minor DNA variations in rice somaclonal variants

Some somaclonal variants derived from a landrace rice variety, Indrayani, were shown to be high yielding and resistant to multiple diseases in previous analysis carried out in our laboratory. An attempt was made to assess the effect of culturing and regeneration of rice plants on DNA variation at microsatellite loci in R2 progeny of callus-derived rice plants. Different somaclones of the rice line Indrayani differing in yield and disease response (high, low and no change in yield, as compared to the original genotype) were used as genetic material for these analyses. Analysis of microsatellite loci was accomplished by digesting DNA from regenerated rice somaclones and assaying for polymorphisms at microsatellite loci by in-gel hybridization with synthetic oligonucleotide probes such as (GATA)4, (CAC)5 and (TG)10. Specific variation at a PCR-amplified locus containing three internal microsatellite repeats (1E6) using restriction site fingerprinting was also investigated. The locus-specific amplification of a sequence-tagged microsatellite marker followed by digestion with HinfI and Sau3AI restriction endonucleases showed differences in some somaclonal variants. The technique used in this study enables monitoring of DNA changes in successive generations of somaclonal variants as a measure of DNA variability and possibly to identify the regions which are responsible for specific traits.

Effect of seasonal acclimatization on the expression of the carp transcription factor Pit-1.

We isolated a clone comprising four exons of the carp Pit-1 gene. Using synthetic oligonucleotide probes derived from the carp Pit-1 sequence Pit-1 expression was assessed by in situ hybridization in pituitary sections from summer- and winter-acclimatized carp. Semiquantitative analyses of the hybridization signals revealed a significant higher Pit-1 expression in the proximal pars distalis (PPD) and pars intermedia (PI) of the pituitary glands from summer-acclimatized carp, compared to the winter-acclimatized fish. In both adaptive states, relative to the PPD and PI, only a basal Pit-1 expression was detected in the rostral pars distalis. Thus, during seasonal acclimatization of an eurythermal fish, Pit-1 seems to be involved in the mechanisms that underlie the compensatory response.

Interleukin-12 and perforin mRNA expression is augmented in blood mononuclear cells in multiple sclerosis.

Cytokines are suggested to orchestrate an abnormal immune response in multiple sclerosis (MS). The regulatory cytokine interleukin (IL)-12 induces T-helper (Th) cell switch to the Th1 type and the production by cytotoxic T cells of perforin, a cell lysis-inducing factor. It has been suggested that Th1-like cytokines may promote the development of MS, and the production of perforin to induce oligodendrocyte damage. In-situ hybridization with radiolabelled synthetic oligonucleotide probes was used to detect and enumerate mononuclear cells (MNC) expressing IL-12 and perforin mRNA in blood and cerebrospinal fluid (CSF) from patients with MS and controls. Plasma and CSF levels of IL-12 (p70) were evaluated by ELISA. Higher numbers of IL-12 and perforin mRNA-expressing MNC were registered in blood in MS and also in controls with aseptic meningoencephalitis (AM) compared to healthy subjects. There were a few patients with other non-inflammatory neurological diseases who also had high levels of IL-12 or perforin mRNA expressing blood MNC. A parallel elevation was observed for IL-12 (p70) concentrations in plasma. In the MS patients' CSF, there was a further augmentation of IL-12 mRNA expressing MNC. To evaluate autoantigen-induced IL-12 and perforin mRNA expression, blood MNC were cultivated +/- myelin basic protein (MBP), a candidate autoantigen in MS. Higher numbers of MBP-reactive IL-12 and perforin mRNA expressing blood MNC were detected in MS than controls. The augmentation of both IL-12 and perforin in MS might reflect ongoing inflammatory processes in MS and could represent targets for future treatments.

Influence of IFN-beta1b (Betaferon) on cytokine mRNA profiles in blood mononuclear cells and plasma levels of soluble VCAM-1 in multiple sclerosis.

Inflammatory cell infiltration within the central nervous system (CNS) and upregulation of both pro- and anti-inflammatory cytokines are characteristic for multiple sclerosis (MS). Treatment with interferon-beta 1b (IFN-beta1b) reduces the number and severity of MS relapses. To examine whether treatment with IFN-beta1b affects levels of cytokine mRNA expressing blood mononuclear cells (MNC) we employed in-situ hybridization with synthetic oligonucleotide probes to detect and enumerate IFN-gamma, TNF-alpha, IL-10, TGF-beta and perforin mRNA expressing cells in MS patients before treatment with IFN-beta1b and during treatment for 3-6 weeks and for 3-6 months. Numbers of blood MNC spontaneously expressing TNF-alpha and IL-10 mRNA were lower after 3-6 months of treatment, while numbers of IFN-gamma, TGF-beta and perforin mRNA expressing MNC were not affected by treatment. IFN-beta1b had no influence on levels of MBP-reactive IFN-gamma, TNF-alpha, TGF-beta, IL-10 or perforin mRNA expressing blood MNC determined after 3-6 weeks or 3-6 months of treatment. Parallel measurements of plasma concentrations of soluble vascular cell adhesion molecule-1 (sVCAM-1) revealed elevated levels after 3-6 weeks of treatment and these levels remained higher after 3-6 months of treatment. The results suggest that IFN-beta1b treatment upregulates plasma levels of sVCAM-1, but has little effects on numbers of blood MNC expressing mRNA of the pro- and anti-inflammatory cytokines under study.Copyright Lippincott-Raven Publishers

A mixture of synthetic oligonucleotide probes labeled with biotin for the sensitive detection of potato spindle tuber viroid

Five kinds of synthetic oligonucleotide probes labeled with biotin (BIO) were designed for the detection of potato spindle tuber viroid (PSTVd), and their sensitivities were compared with that of a digoxigenin (DIG)- or BIO-labeled cDNA probe. Although each oligonucleotide probe alone was less sensitive than the DIG-cDNA probe, sensitivity was increased by using a mixture of two or all of the five oligonucleotide probes. The sensitivity of a PSmix1-5 probe, which was a mixture of five oligonucleotides, was the same as that of a DIG-labeled cDNA probe, which can detect 7.8 pg of purified PSTVd and PSTVd in nucleic acids, equivalent to extracts from 20 μg of infected potato leaf and 310 μg of infected potato tuber. Using the PSmix1-5 probe, PSTVd in all leaves and tubers of seven potato cultivars could be detected without any background. Moreover, with the PSmix1-5 probe, the hybridization time could be shortened to 2 h without any decrease in sensitivity, whereas the sensitivity of the cDNA probes clearly decreased when the hybridization time was shortened. Hybridization using a mixture of several oligonucleotide probes may be applicable to the gene diagnosis of other viroids and viruses.

Ischemic stroke is associated with a systemic increase of blood mononuclear cells expressing interleukin-8 mRNA.

Ischemic brain injury secondary to an arterial occlusion is characterized by acute local inflammation. Blood polymorphonuclear leukocytes (PMNL), primarily neutrophils, adhere to endothelial cells and rapidly invade the injured brain after the arterial occlusion. This neutrophilic invasion might correlate with the production of certain chemoattractants by blood mononuclear cells (MNC). We evaluated mRNA expression of the CXC chemokine interleukin (IL)-8, and the CC chemokines monocyte chemoattractant protein-1, macrophage inflammatory protein (MIP)-1alpha, and MIP-1beta in blood MNC from patients with ischemic stroke. Peripheral blood was obtained at 8 AM on days 1 to 7 (mean, day 3) after onset of symptoms. In situ hybridization with radiolabeled synthetic oligonucleotide probes for the chemokines was adopted to measure chemokine mRNA expression in MNC. An enzyme-linked immunosorbent assay for IL-8 was used to measure IL-8 levels in plasma. Most patients with ischemic stroke had high numbers of IL-8 mRNA expressing blood MNC, regardless of the time interval between onset of clinical symptoms and examination. There was a marked difference between patients with ischemic stroke and healthy subjects (median, 6228 versus 885 positive cells per 10(5) MNC; P<.0001). IL-8 levels in plasma correlated positively to IL-8 mRNA expression in examined patients (n=7) with ischemic stroke (r=.78, P<.05). In contrast, mRNA expression for the CC chemokines showed no significant difference between patients with ischemic stroke and healthy control subjects. This study demonstrated a systemic increase of IL-8 mRNA expressing MNC and IL-8 levels in plasma from patients with ischemic stroke, suggesting that IL-8 could be involved in recruiting blood PMNL to the sites of cerebral ischemia.

Localization of Cortisol Receptor in Branchial Chloride Cells in Chum Salmon Fry

To clarify the involvement of cortisol in functional differentiation of branchial chloride cells, cellular gene expression and localization of cortisol receptor were examined in chum salmon (Oncorhynchus keta) fry in freshwater (FW) and those adapted to seawater (SW) byin situhybridization and immunocytochemical staining. Sodium–potassium adenosinetriphosphatase (Na+,K+-ATPase) activity in the whole gill homogenate was significantly higher in SW fish than in FW fish. There were no significant differences in plasma cortisol levels nor in the expression of cortisol receptor mRNA, examined by Northern blot analysis, between SW and FW fish. The receptor gene expression, examined byin situhybridization with biotin-labeled synthetic oligonucleotide probe, and specific immunostaining with anti-cortisol receptor serum were found in two types of chloride cells distributed in the gill filaments and lamellae, which were also labeled with anti-Na+,K+-ATPase serum, indicating that cortisol may be one of the important factors regulating chloride cell functions. Gene expression of cortisol receptor in filament chloride cells, which were highly activated in SW-adapted fry, was significantly greater in the fry adapted to SW than in FW-adapted fry, reflecting their specific role in salt secretion in SW. Cortisol receptors were also present in undifferentiated cells in the interlamellar regions adjacent to the central venous sinus, also suggesting the involvement of cortisol in the functional differentiation of chloride cells.

Structure of the gene encoding laminaripentaose-producing β-1,3-glucanase (LPHase) of Streptomyces matensis DIC-108

Streptomyces matensis DIC-108 secretes a laminaripentaose-producing β-1,3-glucanase (LPHase) into the culture medium. LPHase was partially purified and its N-terminal amino acid sequence was determined. The gene ( lph) for LPHase was cloned using a colony hybridization technique and a synthetic oligonucleotide probe designed from the N-terminal amino acid sequence, and the nucleotide sequence of lph was determined. lph has an open reading frame of 1203 bp that encode a polypeptide of 401 amino acids including a putative 35-amino acid signal sequence. The mature portion of the polypeptide showed significant sequence similarity with the catalytic domains of β-1,3-glucanases of Oerskovia xanthineolytica and Arthrobacter sp. strain YCWD3. Since lph was not expressed in Escherichia coli, the coding region of lph was amplified by PCR and ligated into the expression vector pKK233-2. E. coli JM109 cells carrying the resultant plasmid pKK- lph produced β-1,3-glucanase of a size identical to that of LPHase produced by S. matensis DIC-108, and the enzyme exclusively liberated laminaripentaose from insoluble β-1,3-glucan.

Characterization of Transcriptional Regulatory Elements in the Promoter Region of the Murine Blood Coagulation Factor VII Gene

To identify the 5' sequences of the murine coagulation factor VII (fVII) gene that resulted in its efficient transcription, a variety of 5'-flanking sequences up to 7 kilobase pairs upstream of the translation ATG initiation codon were fused to the reporter gene, bacterial chloramphenicol acetyltransferase, and relative expression levels of this gene in mouse Hepa 1-6 cells were determined. It was found that the 5' region extending approximately 85 base pairs (bp) upstream of the transcriptional initiation site served as the minimal DNA region that provided full relative promoter activity for chloramphenicol acetyltransferase expression. This region of the gene also contains consensus sequences for liver-enriched transcription factors, C/EBP beta and HNF4, as well as for the ubiquitous protein factors, AP1, H4TF1, NF1, and Sp1. In vitro DNase I footprinting of the 200-bp proximal region of the promoter with a murine Hepa 1-6 cell nuclear extract revealed a clear footprint of a region corresponding to -80 to -28 bp of the murine fVII gene, suggesting that liver factors interact with this region of the DNA. Competitive gel shift and supershift assays with different synthetic oligonucleotide probes demonstrate that proteins contained in the nuclear extract, identified as C/EBP beta, H4TF1, and HNF4, bind to a region of the murine fVII DNA from 85 to 32 bp upstream of the transcription start site. Purified Sp1 also interacts with this region of the DNA at a site that substantially overlaps, but is not identical to, the H4TF1 binding locus. Binding of Sp1 to the mouse DNA was not observed with the nuclear extract as the source of the transcription factors, suggesting that Sp1 is likely displaced from its binding site by H4TF1 in the crude extract. In vivo dimethyl sulfate footprint analysis confirmed the existence of these sites and additionally revealed two other binding regions slightly upstream of the CCAAT/enhancer-binding protein (C/EBP) binding locus that are homologous to NF1 binding sequences. The data demonstrate that appropriate transcription factor binding sites exist in the proximal promoter region of the murine fVII gene that are consistent with its strong liver-based expression in a highly regulated manner.

Isolation and characterization of microsatellite loci in Swietenia humilis (Meliaceae): an endangered tropical hardwood species

AbstractMicrosatellites are now firmly established as an informative marker system, with increasing popularity as a tool amongst molecular ecologists. We have developed a method of constructing an enriched microsatellite library for the tropical tree species Swietenia humilis Zucc. (Meliaceae). This method is based on a precloning enrichment of SSRs using synthetic oligonucleotide probes, bound to magnetic beads and hybridizing to complementary microsatellite core sequences in digested genomic DNA. Here we describe the isolation and characterization of 10 microsatellite loci that have been used to survey the genetic diversity within a natural population of S. humilis. A total of 97 alleles were identified with an average of 9.7 alleles over all loci. Very high levels of allelic polymorphism were detected at individual loci, with 23 alleles observed at the most variable. The mean observed heterozygosity was 0.415 (range 0.038‐0.815) exceeding levels of diversity detected in related species which used isozymes as the marker system. Subpopulation differentiation at a microgeographical scale was low (FST= 0.036) and the values of Nm, calculated from the allelic frequencies, were greater than 1 thus reflecting the extent of gene flow occurring between individual trees.

Autoantigen‐induced IL‐13 mRNA expression is increased in blood mononuclear cells in myasthenia gravis and multiple sclerosis

Evidence has been presented for the involvement of immune mechanisms in the pathogenesis of myasthenia gravis (MG) and multiple sclerosis (MS). The production of autoantibodies in both diseases is regulated by T‐cells by means of cytokines. Interleukin‐13 (IL‐13) is mainly produced by T‐helper type 2 cells and induces B‐cell proliferation and antibody class switch. The role of IL‐13 in MG and MS is not known. We employed in situ hybridization with synthetic radiolabelled oligonucleotide probes to detect and enumerate blood and cerebrospinal fluid (CSF) mononuclear cells (MNC) expressing IL‐13 mRNA from patients with MG, MS, optic neuritis (ON), other inflammatory neurological diseases (OIND) and healthy controls. MG is associated with elevated levels of acetylcholine receptor (AChR) reactive IL‐13 mRNA expressing blood MNC compared to control patients. In MS, numbers of MBP‐reactive IL‐13 mRNA expressing MNC were higher compared to cultures without antigen stimulation. The levels of MBP‐reactive IL‐13 mRNA positive MNC were higher in MS compared to MG, but not other controls. There were no differences in spontaneous IL‐13 mRNA expressing blood MNC numbers between MG, MS, ON and control patients. The data suggest the involvement of IL‐13 in both MG and MS.

Expression of LAZ3/BCL6 in follicular center (FC) B cells of reactive lymph nodes and FC-derived non-Hodgkin lymphomas.

Chromosomal translocation resulting in abnormal expression of the LAZ3/BCL6 gene in B cells has been implicated in the tumorigenesis of non-Hodgkin lymphoma (NHL). Therefore we studied the expression pattern of LAZ3/BCL6 by in situ hybridization with synthetic oligonucleotide probes in frozen tissue sections from five reactive lymph nodes and 38 B cell and non-B NHL. In addition, we investigated the expression of LAZ3/BCL6 by Northern blot analysis on multiple human tissues. The LAZ3/BCL6 transcript was found in a variety of tissues, including skeletal muscle, peripheral blood leukocytes, and weakly in normal lymph nodes. In the tumor samples, expression of LAZ3/BCL6 was observed in 68% of all B cell NHL and none of the non-B lymphomas. All cases of follicular, mixed small and large cell lymphomas showed LAZ3/BCL6 expression confined to the neoplastic follicles. A follicular expression pattern was also found in all non-malignant reactive lymph nodes. Hence, the expression of LAZ3/BCL6 does not correlate to malignancy, but reflects the origin of B cells from the germinal centers.

Effect of Temperature and Photoperiod on Prolactin Transcription in Cyprinus carpio

Abstract We studied the effect of temperature and photoperiod on prolactin (PRL) gene expression in the pituitaries of summer-aclimatized carp (C. carpio). To assess the level of PRL gene transcription, we used a 24mer synthetic oligonucleotide probe derived from the known carp PRL gene sequence. Employing in situ hybridization assays, we observed high expression of PRL mRNA in the rostral pars distalis of summer-acclimatized carp in contrast to the almost negligible PRL transcription which occurs in the winter-acclimatized fish. When experimental combinations of long and short photoperiods with 10°C and 20°C environmental temperatures were studied, only a short photoperiod (8L-16D) in summer-acclimatized carp acclimated to a winter temperature (10°C) markedly depressed PRL gene expression. Our observations indicate that photoperiod constitutes a particularly relevant modulator in the neuroendocrine cascade that activates PRL transcription in the carp.

Synthetic DNA-based genetic markers reveal intra- and inter-species DNA sequence variability in the Bubalus bubalis and related genomes.

A series of synthetic oligonucleotide probes were developed as markers for genetic analysis and molecular systematics of Bubalus bubalis and other eutherian mammals. A very high level of heterozygosity (approximately 95%) was observed in the bubaline genome, with an overall band-sharing probability of 2.08 x 10(-11) using (TGG)6 and Hinf I probe-enzyme combination. Breed affiliation studies on different buffalo breeds, viz. Toda, Surti, Mehsana, and Murrah, revealed that the semiwild Toda breed belonged to a distinct group. The desired genotypes in each successive generation in an actual breeding program were also identified by employing a DNA fingerprinting approach using these markers. Estimation of genetic distances by calculating the mean allelic frequencies at (CA)n, (TGG)n, and (GGAT)n repeat loci between buffalo and other related animals such as horse (order Perisodactyla), rabbits (order Lagomorpha, (pigs, cattle, goat, and sheep (order Artiodactyla) revealed that with respect to the genetic distance coefficient (GDC), goat (caprine) was closer to buffalo (GDC = 0.0005) than sheep (ovine) (GDC = 0.0007). The genetic distance between horse and buffalo was calculated to be 0.4085, indicating that compared to other animals, horse was distantly related to buffalo. The understanding of overall allelic variations and breed affiliation of the bubaline genome will contribute to the propagation and conservation of the desired germ plasm and better management of this species.

Genetic determination of paternity and variation in male reproductive success in two populations of rhesus macaques.

The fundamental framework for uncovering factors affecting the evolution of social behavior rests upon analyses of variation in reproductive success. In species where females mate with multiple males, paternity is invisible in the absence of genetic data. We determined paternity in two populations of rhesus macaques, Macaca mulatta, using both single locus and multilocus techniques. One troop, Group R, is one of four troops living on a 15 ha island (Cayo Santiago) off the coast of Puerto Rico, while the other troop, Group M, was translocated from Cayo Santiago to the Sabana Seca Field Station (Puerto Rico) in 1984. About a dozen human-derived short tandem repeat (STR) markers have been found to be polymorphic in the study of populations and provide the initial paternity determination. Final evaluation of paternity is then confirmed by multilocus DNA fingerprinting using synthetic oligonucleotide probes. Body condition, age, and dominance rank have an impact on male progeny production, while canine size does not. We suggest that nonagonistic competition in the form of sperm competition and endurance rivalry will modulate male reproductive success. A large body size among males provides them with an advantage in both sperm competition and endurance rivalry. Comparison of the two populations indicated that demographic, social, ecological, and morphological factors interact to regulate variation in reproductive success among male nonhuman primates.

Cloning and Characterization of a Novel Oxidoreductase KDRF from a Human Bone Marrow-derived Stromal Cell Line KM-102

A cDNA clone coding for a novel oxidoreductase was cloned from a human bone marrow-derived stromal cell line KM-102. We screened a cDNA library constructed from the mRNA of KM-102 cells stimulated with phorbol 12-myristate 13-acetate and calcium ionophore A23187 using a 32P-labeled 15-mer synthetic oligonucleotide (5'-TAAATAAATAAATAA-3') probe. This probe was designed as a complementary sequence to the three reiterated AUUUA sequences, which are contained in the 3'-untranslated regions of cytokine and some proto-oncogene mRNAs and correlate with rapid mRNA turnover. Then, we obtained one cDNA clone, and further sequence analysis revealed that it coded for a new protein exhibiting 30 to approximately 40% homology with glutathione reductase. By fusion protein analysis, this protein showed reducing activities on 2, 6-dichlorophenol-indophenol and 5,5'-dithio-bis(2-nitrobenzoic acid) but only a weak reducing activity on oxidized glutathione. Although it lacked a stretch of hydrophobic amino acids in its N terminus, it was secreted by monkey kidney-derived COS-1 cells when we introduced the expression plasmid into them and also secreted by a human lung carcinoma cell line A549. Northern blot analysis revealed that the mRNA turnover of this protein was regulated by inflammatory stimuli in KM-102 cells. These results show that this protein may have scavenging enzyme properties and has its mRNA expression regulated in a similar fashion to cytokine genes or proto-oncogenes. Thus, we named it KDRF (KM-102-derived reductase-like factor), and KDRF may play a role in scavenging reactive oxygen intermediates, which are possibly toxic to cells, in response to inflammatory stimuli.