Expression Of TR3 Research Articles

Dihydromyricetin from ampelopsis grossedentata protects against vascular neointimal formation via induction of TR3

Vine tea has been used as a medicinal herb in traditional Chinese medicine for hundreds of years. As the most abundant ingredient in vine tea, Dihydromyricetin (DHM) has been reported to exert anti-inflammatory, antioxidant, and anti-cardiovascular disease. However, the role of DHM in injury-induced neointimal formation remains poorly characterized. We determined the effects of DHM on ligation-induced carotid artery neointimal formation. We found that ligation-induced carotid artery neointimal formation could be significantly attenuated by DHM treatment. We provide evidence that DHM increases orphan nuclear receptor TR3 expression in smooth muscle cell (SMC) and carotid artery. Moreover, overexpression and loss-of-function strategies of TR3 were done to overexpression and knockdown of TR3, and demonstrate that DHM promotes SMC differentiation, however, inhibits SMC proliferation and migration, via regulating expression of TR3. Collectively, we reveal that DHM may be a therapeutic agent for the treatment of injury-induced vascular diseases.

Orphan Nuclear Receptor TR3/Nur77 is a Specific Therapeutic Target for Hepatic Cancers.

Although great success has been achieved in cancer treatment, current cancer therapies, including anti-tumorigenesis and anti-angiogenesis, still face the problems of insufficient efficacy, resistance and intrinsic refractoriness, in addition to their toxic side effects. There is a demand to identify additional targets that can be blocked to turn off the downstream effects of most, if not all, pathways. Our previous studies suggest that orphan nuclear receptor TR3 (human) / Nur77 (mouse) is such a target. However, the correlation of TR3 expression and clinical tumor progression has not been studied. The expression of TR3 was analysed in human primary hepatic cancer specimens from patients that have complete medical records with Immunohistochemical staining. The statistical analysis was used to assess the significance of TR3 expression in tumor tissues, paratumor tissues and normal tissues, and to investigate the correlation of TR3 expression and clincopathologic characteristics. TR3 is highly expressed in human hepatic cancer tissues, but not in normal liver tissues. The positive expression yields of TR3 are 67.67% (14/21), 19.05% (4/21) and 0% (0/10) in cancer tissues, para cancer tissues, and normal liver tissue, respectively, which are statistic significant (χ2=17.07, p<0.005). The expression of TR3 is significantly higher in cancer tissues than in para cancer tissues χ2=9.722, p<0.005) and in normal tissues (p<0.0005). The levels of TR3 expression in human hepatic cancer tissues correlates well with tumors that are at low/middle degree of tumor differentiation and have portal vein thrombosis, metastasis and recurrence, but not with age, gender, tumor number and Alpha-fetal protein (AFP) volume. The results indicate that TR3 is a specific therapeutic target for hepatic cancers.

TR3 is preferentially expressed by bulge epithelial stem cells in human hair follicles

TR3 is an orphan member of the steroid/thyroid/retinoid nuclear receptor superfamily of transcription factors and it plays a pivotal role in regulating cell growth and apoptosis. The expression and function of TR3 in skin have not been well investigated. Using a cDNA expression assay, we discover that TR3 is significantly enriched in human telogen bulge compared with anagen bulb. Immunohistochemical staining confirms that TR3 is highly expressed in the bulge region of human hair follicles and it colocalizes with cytokeratin 15 (K15), an epithelial stem cell marker. To study the function of TR3 in the effect of androgens in keratinocytes, we treat HaCaT keratinocytes and primary human keratinocytes with dihydrotestosterone (DHT) and testosterone (T). The treated keratinocytes show a dose-dependent growth reduction to DHT and T. DHT increases the expression of TR3 in keratinocytes, associated with a concomitant increase of BAD and decrease of Bcl-2 expression. Knockdown TR3 expression by siRNA blocks the inhibitory effect of DHT on keratinocyte proliferation. Our results demonstrate that TR3 is localized to the stem cell compartment in the human hair follicles. Androgen increases TR3 expression in cultured keratinocytes. Our data suggest that TR3 mediates at least part of the inhibitory effect of androgens on keratinocytes.

Differential regulation of orphan nuclear receptor TR3 transcript variants by novel vascular growth factor signaling pathways.

Angiogenesis is a hallmark of many diseases, including cancer, ischemic heart disease, inflammation, and others. It is well known that vascular endothelial growth factor (VEGF) is the most important angiogenic factor. Recently, we demonstrated that orphan nuclear receptor TR3 (mouse Nur77 and rat NGFI-B) plays critical roles in tumor growth and angiogenesis induced by VEGF-A in vitro and in vivo. However, the signaling pathways that mediate the expression of TR3 induced by VEGF are still not completely understood. Here we reported that 3 TR3 transcript variants (TR3-TVs) are expressed at differential levels, and regulated differentially in endothelial cells. While the expression of TR3-TV1 is relatively low, the expression of TR3-TV2 is up-regulated markedly, and the expression of TR3-TV3 is up-regulated moderately in endothelial cells induced by VEGF-A. The kinetics of the induction of these TR3-TVs is different. We also found that several signaling pathways, including calcium-PLC-PKC-PKD1 pathway, NF-κB pathway, and MAP kinase (ERK, p38, and JNK) pathways are important for VEGF-A-induced TR3-TV2 and TR3-TV3 mRNA induction. More important, we found that VEGF-A or VEGF-E, but not VEGF-B, nor placenta growth factor (PlGF), induces the phosphorylation of insulin-like growth factor-1 receptor (IGF-1R) and the interaction of VEGF receptor 2/kinase insert domain receptor (VEGFR2/KDR) with IGF-1R, which mediates the expression of TR3-TV2, but not TR3-TV3. Taking together, we demonstrate that TR3-TVs are differentially regulated by VEGF-A and identify a novel signaling pathway by which VEGF-A and VEGF-E, but neither VEGF-B, nor PlGF, induce the interaction of VEGFR2/KDR with IGF-1R, resulting in IGF-1R transactivation to induce the high level expression of TR3-TV2. Our data not only elucidate the signaling pathways by which TR3-TVs are regulated, but extend the molecular mechanism, by which VEGF-A-induced angiogenesis. These studies should permit the development of screening assays for compounds that inhibit VEGF signaling.

Inactivation of the nuclear receptor TR3 as an important new drug target for treating RCC.

e15618 Background: The high mortality of patients with RCC is due to poor performance of most chemotherapy. There is a critical need for development of new mechanism-based drugs.The nuclear orphan receptor NR4A1 (TR3, Nur77) are immediate early genes activated by various stressors. NR4A is highly expressed in multiple tumors. TR3 knockdown studies show that TR3 plays an important role in cancer cell growth and angiogenesis. The pro-oncogenic activity of this receptor is an excellent example of “non-oncogene addiction” which is essential for maintaining the cancer cell/tumor phenotype. Methods: Multiple RCC cell lines have been used. Several different classes of C-substituted DIMs summarized in Fig1 has been screened for TR3 deactivation in RCC cell lines using GAL4-TR3/UAS-luc constructs (Fig1). Effects of the C-DIM compounds on cancer cell growth and expression of Sp-regulated genes has been determined by cell counting, the MTT assay, western blot and activation of caspases 3 and 9 and directly compared to knockdown of TR3 by RNAi. Results: Knockdown of TR3 by RNAi has confirmed that TR3 is pro-oncogenic and we have focused on structure-activity relations (SARs) of C-DIM analogs to identify compounds that inactivate TR3. Renal cancer cells were treated with c-DIM TR3 inactivators and this significantly inhibited cell proliferation (Figs 2 and 3). Expression of TR3 in RCC cell lines has not previously been reported. Fig. 4 presents a western blot of cell lysates showing that TR3 is expressed in these cell lines. TR3 inactivators decreased expression of survivin and bcl-2 and this was accompanied by several apoptotic markers. These results show that RCC cells not only express TR3 but TR3 inactivators decrease the pro-survival genes and induce apoptosis which confirm that TR3 is also a target for renal cancer therapy. TR3 pro-oncogenic activity is due, in part, to the role of TR3 in constitutive expression of survivin and other Sp-regulated genes (VEGF, c-MET..). TR3 also involves in formation of a complex with p53 which inactivates the tumor suppressor gene. Thus inactivation of TR3 activates p53 pathway (Fig 5). Conclusions: TR3 is overexpressed in RCC. We have shown for the first time that TR3 is an important new drug target for treating RCC.

The orphan receptor TR3 participates in angiotensin II‐induced cardiac hypertrophy by controlling mTOR signalling

Angiotensin II (AngII) induces cardiac hypertrophy and increases the expression of TR3. To determine whether TR3 is involved in the regulation of the pathological cardiac hypertrophy induced by AngII, we established mouse and rat hypertrophy models using chronic AngII administration. Our results reveal that a deficiency of TR3 in mice or the knockdown of TR3 in the left ventricle of rats attenuated AngII-induced cardiac hypertrophy compared with the respective controls. A mechanistic analysis demonstrates that the TR3-mediated activation of mTORC1 is associated with AngII-induced cardiac hypertrophy. TR3 was shown to form a trimer with the TSC1/TSC2 complex that specifically promoted TSC2 degradation via a proteasome/ubiquitination pathway. As a result, mTORC1, but not mTORC2, was activated; this was accompanied by increased protein synthesis, enhanced production of reactive oxygen species and enlarged cell size, thereby resulting in cardiac hypertrophy. This study demonstrates that TR3 positively regulates cardiac hypertrophy by influencing the effect of AngII on the mTOR pathway. The elimination or reduction of TR3 may reduce cardiac hypertrophy; therefore, TR3 is a potential target for clinical therapy.

Nuclear orphan receptor TR3/Nur77 mediates melanoma cell apoptosis

TR3 was originally recognized for its role in the regulation of cell survival and differentiation, however, it was recently found to be a potent pro-apoptotic protein. In order to characterize the role of TR3 in melanoma cell apoptosis, we studied expression of TR3 in melanoma cell lines and tissues, its subcellular distribution and function during apoptosis using various expression and RNA interference vectors. We found that TR3 was constitutively expressed in both cultured melanoma cells and melanoma tissues. TR3 expression was significantly decreased in advanced melanomas comparing to benign nevi. Over-expression of wild type TR3 or mutant TR3 lacking the DNA binding domain resulted in massive apoptosis in melanoma cells, whereas stable knockdown of TR3 using RNA interference resulted in melanoma cell resistance to apoptosis induced by chemotherapeutic agents ATRA and cisplatin. We further demonstrated that apoptosis in melanoma cells was mediated, at least partially, through TR3 mitochondrial translocation but not alteration in TR3 expression levels. Our results suggest that TR3 is an important apoptosis inducing factor in melanoma cells. Decreased expression of TR3 in metastatic melanoma cells may contribute to their reduced apoptotic potential and increased resistance to chemotherapy.

Expression of Orphan Receptors TR2, TR3, TR4, and p53 in Heat‐Treated Testis of Cynomolgus Monkeys (Macaca fascicularis)

To investigate the possible role of testicular orphan receptors (TR) TR2, TR3, and TR4 in the process of germ cell apoptosis in the heat-treated testis of monkey, we have examined the spatiotemporal expression of the 3 TR mRNAs in relation to p53 mRNA levels in the monkey testis by in situ hybridization and reverse transcription polymerase chain reaction techniques. The results showed that TR2 mRNA was confined to spermatocytes; TR4 and TR3 mRNAs were expressed in both spermatocytes and spermatids. The heat treatment did not change TR2 mRNA level but significantly reduced TR4 mRNA expression in spermatocytes on days 3 and 8 after the heat treatment. TR3 mRNA expression was affected by the heat treatment in a time-dependent manner, with the lowest level on day 30 after the heat shock. Low to moderate signal for p53 mRNA was detected in spermatocytes before treatment, which increased dramatically on days 3, 8, and 30 after the heat shock. The coincident expression of the testicular TR3 and p53 mRNA, spatially and time dependently, implied that the decrease in TR3 expression in the heat-treated testis might be closely related to the p53 signal pathway, whereas the temporal decrease in TR4 production in the testis at the early stage indicated that this orphan receptor might be also involved in germ cell apoptosis. The data suggest that TR3, TR4, and p53 could be important regulators of germ cell apoptosis induced by the heat treatment, whereas TR2 might not be a key regulator in this process.

Orphan nuclear receptor TR3/Nur77 regulates VEGF-A–induced angiogenesis through its transcriptional activity

Vascular endothelial growth factor (VEGF)-A has essential roles in vasculogenesis and angiogenesis, but the downstream steps and mechanisms by which human VEGF-A acts are incompletely understood. We report here that human VEGF-A exerts much of its angiogenic activity by up-regulating the expression of TR3 (mouse homologue Nur77), an immediate-early response gene and orphan nuclear receptor transcription factor previously implicated in tumor cell, lymphocyte, and neuronal growth and apoptosis. Overexpression of TR3 in human umbilical vein endothelial cells (HUVECs) resulted in VEGF-A–independent proliferation, survival, and induction of several cell cycle genes, whereas expression of antisense TR3 abrogated the response to VEGF-A in these assays and also inhibited tube formation. Nur77 was highly expressed in several types of VEGF-A–dependent pathological angiogenesis in vivo. Also, using a novel endothelial cell-selective retroviral targeting system, overexpression of Nur77 DNA potently induced angiogenesis in the absence of exogenous VEGF-A, whereas Nur77 antisense strongly inhibited VEGF-A–induced angiogenesis. B16F1 melanoma growth and angiogenesis were greatly inhibited in Nur77−/− mice. Mechanistic studies with TR3/Nur77 mutants revealed that TR3/Nur77 exerted most of its effects on cultured HUVECs and its pro-angiogenic effects in vivo, through its transactivation and DNA binding domains (i.e., through transcriptional activity).

Tcl-1 Inhibits Nuclear Export of TR3 and Is Highly Expressed in ZAP70-Positive CLL B-Cells.

Chronic Lymphocytic Leukemia (CLL) is the most frequent leukemia afflicting the Western world. Unlike other more aggressive leukemias and lymphomas, CLL B-cells do not divide at a high rate and expansion of the malignant clone appears to be due to underlying defect in apoptosis.Elevated levels of anti-apoptotic Bcl-2 are thought to be involved in the pathogenesis of CLL. The activity of Bcl-2 can be modulated my interactions with other cellular proteins. Recently, orphan nuclear receptor TR3 (nur77) was shown to exit the nucleus, bind Bcl-2, and convert Bcl-2 into a killer protein (Lin, et al., CELL , 116:527, 2004). We evaluated the expression of TR3 by immunoblotting in CLL B-cells specimens. High levels of TR3 protein were present in 8 of 8 CLL patient specimens. Subcellular fractionation studies showed predominately nuclear localization of TR3 in 2 of 2 CLL specimens examined. Because Akt is known to regulate TR3 activity in the nucleus (Pekarsky, et al., Proc. Natl. Acad. Sci. 98:3690, 2001), we also examined the expression of the Akt regulator, Tcl-1. The Tcl-1 protein was expressed in the majority of CLL samples and at high levels in 8 of 16 CLL specimens, correlating with ZAP70-positivity. In gene transfection experiments using tumor cell lines, over-expression of Tcl-1 inhibited nuclear export of TR3, as determined by immunofluorescence confocal microscopy. We propose a model in which expression of the oncogene Tcl-1 in CLL B-cells (perhaps related to ZAP70 expression) modulates nuclear Akt activity, in turn suppressing TR3 function, and thus reducing activity of the TR3-mediated pathway for conversion of Bcl-2 from protector to killer. This hypothesis requires further experimental testing.

Mitogenic effect of orphan receptor TR3 and its regulation by MEKK1 in lung cancer cells.

TR3, also known as NGFI-B or nur77, is an immediate-early response gene and an orphan member of the steroid/thyroid/retinoid receptor superfamily. We previously reported that TR3 expression was induced by apoptotic stimuli and was required for their apoptotic effect in lung cancer cells. Here, we present evidence that TR3 was also induced by epidermal growth factor (EGF) and serum and was required for their mitogenic effect in lung cancer cells. Ectopic expression of TR3 in both H460 and Calu-6 lung cancer cell lines promoted their cell cycle progression and BrdU incorporation, while inhibition of TR3 expression by the small interfering RNA approach suppressed the mitogenic effect of EGF and serum. Analysis of TR3 mutants showed that both TR3 DNA binding and transactivation were required for its mitogenic effect. In contrast, they were dispensable for its apoptotic activity. Furthermore, confocal microscopy analysis demonstrated that TR3 functioned in the nucleus to induce cell proliferation, whereas it acted on mitochondria to induce apoptosis. In examining the signaling that regulates the mitogenic function of TR3, we observed that coexpression of constitutive-active MEKK1 inhibited TR3 transcriptional activity and TR3-induced proliferation. The inhibitory effect of MEKK1 was mediated through activation of Jun N-terminal kinase, which efficiently phosphorylated TR3, resulting in loss of its DNA binding. Together, our results demonstrate that TR3 is capable of inducing both proliferation and apoptosis in the same cells depending on the stimuli and its cellular localization.

TR3 orphan receptor is expressed in vascular endothelial cells and mediates cell cycle arrest.

Endothelial cells play a pivotal role in vascular homeostasis. In this study, we investigated the function of the nerve growth factor-induced protein-B (NGFI-B) subfamily of nuclear receptors comprising the TR3 orphan receptor (TR3), mitogen-induced nuclear orphan receptor (MINOR), and nuclear orphan receptor of T cells (NOT) in endothelial cells. The mRNA expression of TR3, MINOR, and NOT in atherosclerotic lesions was assessed in human vascular specimens. Each of these factors is expressed in smooth muscle cells, as described before, and in subsets of endothelial cells, implicating that they might affect endothelial cell function. Adenoviral overexpression of TR3 in cultured endothelial cells resulted in decreased [3H]thymidine incorporation, whereas a dominant-negative TR3 variant that inhibits the activity of endogenous TR3-like factors enhanced DNA synthesis. TR3 interfered with progression of the cell cycle by upregulating p27Kip1 and downregulating cyclin A, whereas expression levels of a number of other cell cycle-associated proteins remained unchanged. These findings demonstrate that TR3 is a modulator of endothelial cell proliferation and arrests endothelial cells in the G1 phase of the cell cycle by influencing cell cycle protein levels. We hypothesize involvement of TR3 in the maintenance of integrity of the vascular endothelium.

Focusing on transcription factor families in atherogenesis: the function of LKLF and TR3

In this overview, two separate studies are discussed that emerged from a "discovery-driven" approach to identify genes that play an essential role in atherogenesis. First, by a combination of DNA micro-array and one-way linkage hierachical clustering, we selected genes that are induced in endothelial cells (EC) by prolonged steady- or pulsatile laminar flow, but of which expression is not affected by inflammatory and mitogenic agents (TGF-beta, IL-1betaTNF-alpha,VEGF, thrombin). The genes selected accordingly were: cytochrome P450 1B1, diaphorase and the transcription factor lung Krüppel-like factor (LKLF) of which only the latter is truly EC specific. LKLF meets the criteria of an anti-atherosclerotic gene, mainly since expression is restricted to areas subjected to laminar flow as shown by in situ hybridization with anatomically well-defined specimens. Second, neointimal (but not medial) smooth muscle cells (SMC) specifically synthesize the NGFI-B subfamily (TR3, MINOR,NOT) of the nuclear hormone superfamily of transcription factors. Again, evidence is presented, indicating that these proteins serve an anti-atherosclerotic function. Notably, transgenic mice, expressing either TR3 or a dominant-negative mutant TR3DeltaTA in arterial SMC, were subjected to carotid artery ligation to induce SMC proliferation. Lesions in TR3-overexpressing transgenic mice were 5-fold smaller than isogenic wild-type mice, while mice overexpressing the TR3DeltaTA mutant had a 3-fold larger lesion. It is proposed that (down-stream products of) TR3 inhibit the cell cycle, since adenovirus-mediated expression of TR3DeltaTA and TR3, respectively, inhibit and promote the synthesis of the cyclin-dependent kinase inhibitor p27(Kip1) in SMC.

Protective function of transcription factor TR3 orphan receptor in atherogenesis: decreased lesion formation in carotid artery ligation model in TR3 transgenic mice.

Smooth muscle cells (SMCs) play a key role in intimal thickening in atherosclerosis and restenosis. The precise signaling pathways by which the proliferation of SMCs is regulated are largely unknown. The TR3 orphan receptor, the mitogen-induced nuclear orphan receptor (MINOR), and the nuclear receptor of T cells (NOT) are a subfamily of transcription factors belonging to the nuclear receptor superfamily and are induced in activated SMCs. In this study, we investigated the role of these transcription factors in SMC proliferation in atherogenesis. Multiple human vascular specimens at distinct stages of atherosclerosis (lesion types II to V by American Heart Association classification) derived from 14 different individuals were studied for expression of these transcription factors. We observed expression of TR3, MINOR, and NOT in neointimal SMCs, whereas no expression was detected in medial SMCs. Adenovirus-mediated expression of a dominant-negative variant of TR3, which suppresses the transcriptional activity of each subfamily member, increases DNA synthesis and decreases p27(Kip1) protein expression in cultured SMCs. We generated transgenic mice that express this dominant-negative variant or full-length TR3 under control of a vascular SMC-specific promoter. Carotid artery ligation of transgenic mice that express the dominant-negative variant of TR3 in arterial SMCs, compared with lesions formed in wild-type mice, results in a 3-fold increase in neointimal formation, whereas neointimal formation is inhibited 5-fold in transgenic mice expressing full-length TR3. Our results reveal that TR3 and possibly other members of this transcription factor subfamily inhibit vascular lesion formation. These transcription factors could serve as novel targets in the treatment of vascular disease.

Induction of apoptosis by TPA and VP-16 is through translocation of TR3.

To investigate the role of TR3 in induction of apoptosis in gastric cancer cells. Human gastric cancer cell line, MGC80-3, was used. Expression of TR3 mRNA and its protein was detected by Northern blot and Western blot. Localization of TR3 protein was showed by immunofluorescence analysis under laser-scanning confocal microscope. Apoptotic morphology was observed by DAPI fluorescence staining, and apoptotic index was counted among 1000 cells randomly. Stable transfection assay was carried out by Lipofectamine. Treatment of MGC80-3 cells with TPA and VP-16 resulted in apoptosis, accompanied by the repression of Bcl-2 protein in a time-dependent manner. At the same time, TPA and VP-16 also up-regulated expression level of TR3 mRNA in MGC80-3 cells that expressed TR3 mRNA. When antisense-TR3 expression vector was transfected into the cells, expression of TR3 protein was repressed. In this case, TPA and VP-16 did not induce apoptosis. In addition, TPA and VP-16-induced apoptosis involved in translocation of TR3. In MGC80-3 cells, TR3 localized concentrative in nucleus, after treatment of cells with TPA and VP-16, TR3 translocated from nucleus to cytosol obviously. However, when this nuclear translocation was blocked by LMB, apoptosis was not occurred in MGC80-3 cells even in the presence of TPA and VP-16. Induction of apoptosis by TPA and VP-16 is through induction of TR3 expression and translocation of TR3 from nucleus to cytosol, which may be a novel signal pathway for TR3, and represent the new biological function of TR3 to exert its effect on apoptosis in gastric cancer cells.

Localization of orphan receptor TR3 mRNA in early developmental follicles in rat

Byin situ hybridization, the localization of orphan receptor TR3 mRNA has been observed in early developmental follicles. TR3 mRNA is first expressed in the ovarian interstitial cells on day 2 after birth, and then in granulosa cells (GC) in primary follicles on day 4. The expression level of TR3 mRNA in GC increases following the follicular development. Its higher expression can be observed in the outer layer of GC and inner layer of theca cells (TC) on day 6, where the cells present active proliferation and differentiation. The expression of TR3 is in an increasing manner until the large antral follicles on day 30. The mRNA is only expressed in the healthy, but not atretic follicles in adult rat ovaries. Injection of epidermal growth factor (EGF) has dramatically enhanced its expression in the early stage of developmental follicles. It is therefore suggested that TR3 may play a role in regulating growth and differentiation of ovarian somatic cells in the early stage, and its expression is regulated by EGF.

TR3 death receptor expression in the normal and ischaemic brain

Members of the death receptor family may play a prominent role in developmental and pathological neuronal cell death. We report the expression of the TR3 and TR7 death receptors in the adult human and rat central nervous system. Whereas expression of TR3 appears to be high in the human cerebellum, with lower levels in other brain regions, robust expression is observed in many regions of the rat brain. We also analysed modulation of death receptor expression in an in vivo rat model of acute stroke. In contrast to tumour necrosis factor receptor 1, Fas and p75 NGFR, which all show up-regulation specifically in lesioned cortex of the permanent middle cerebral artery occlusion model of stroke, TR3 shows a rapid global increase in both lesioned and unlesioned brain. In comparison, the recently described death receptor TR7 shows no change in this model. These data indicate that the death receptors show clear differences in patterns of expression in response to ischaemic injury.

Characterization of a novel TNF-like ligand and recently described TNF ligand and TNF receptor superfamily genes and their constitutive and inducible expression in hematopoietic and non-hematopoietic cells.

A novel (TL1), a recently described (TL2) TNF-like, and three recently described TNF receptor-like (TR1, TR2, TR3) molecules were identified by searching a cDNA database. TL1 and TL2 are type-II membrane proteins. TR2 and TR3 are type-I membrane proteins whereas TR1 appears to be a secreted protein. TL1, TL2, TR2 and TR3 were expressed in hematopoietic cells, whereas TR1 was not. Northern blots hybridized with the cDNA probes revealed multiple forms of RNA as well as inducible expression of TL1, TL2, TR2 and TR3. TL2 and TR3, in particular, were highly induced in activated CD4+ T cells. Radiation hybrid mapping localized TR1 and TL2 to 8q24 and 3q26, respectively, which are not near any known superfamily members. TL1 was mapped to 9q32, near CD30L (9q33) and TR2 and TR3 mapped to the region of chromosome 1 that contains the TNFR-II, 4-1BB, OX40 and CD30 gene cluster at 1p36. Only TR3 in this cluster possesses a death domain. Southern blot analysis revealed the presence of TL and TR genes in different mammalian species. TL2, TR1, TR2 and TR3 were recently described by others as TRAIL/Apo-2L, OPG, HVEM and DR3/WSL-1/Apo-3/TRAMP/LARD, respectively.