Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Head and neck cancer arises and progresses through specific genetic alterations which lead to an invasive immortal phenotype. The process of immortalization is associated with the activation of the enzyme telomerase, a ribonucleoprotein with reverse transcriptase activity which is capable of synthesizing telomeric repeats at the end of chromosomes. This enzyme is expressed in nearly all neoplasms and germline cells and is absent in most normal human somatic cells. Because of this expression pattern testing for telomerase activity may deliver useful diagnostic and/or prognostic information about clinical tumour behaviour. Telomerase activity was therefore analysed in 16 primary lesions of head and neck squamous cell carcinomas (HNSCC) using the polymerase chain reaction-based telomeric repeat amplification protocol (TRAP). For a sensitive semiquantitative analysis of telomerase activity TRAP products were mixed with Pico Green I and the fluorescence emission intensities were measured. All 16 samples tested positive. When the Pico Green I data were compared with clinical parameters, it was obvious that N0 necks revealed significantly (p < 0.05) lower emission intensities (i.e. telomerase activity) than N + necks. Our results indicate that a high telomerase activity in HNSCC may facilitate lymph node metastasis and that the estimation of telomerase activity is a useful diagnostic tool which could influence treatment modalities.
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PMID:Non-radioactive semiquantitative testing for the expression levels of telomerase activity in head and neck squamous cell carcinomas may be indicative for biological tumour behaviour. 965 21

Long interspersed elements, or LINEs, are retrotransposons that move via an RNA intermediate. In mice, one polymorphic variant of L1 has amplified relatively recently, giving rise to the A-type subfamily in species belonging to the genus and subgenus Mus. Retrotransposition of LINE-1 (L1) requires the function of the L1-encoded reverse transcriptase that is produced from open reading frame 2 (ORF2). Here, we employ a convenient yeast genetic assay to determine the reverse transcriptase activity of the ORF2 obtained from three A-type L1 elements: one, a cDNA from the RNA in ribonucleoprotein particles; another with a purported inactivating mutation; and the third, a hypothetical ancestral construct. Because there are no examples of A-type elements that have transposed recently to inactivate a gene, this assay is the first step towards demonstrating the functional capability of mouse A-type LINE-1 elements. One of the three elements was believed to have been inactivated during evolution by the substitution of leucine for a highly conserved phenylalanine or tryptophan residue among known reverse transcriptases. This mutation did not inactivate the L1 reverse transcriptase in the yeast assay; thus, all three of the elements tested encoded reverse transcriptase activity. We further examined the minimal reverse transcriptase domain within ORF2 by creating a series of deletions. The results demonstrate that removal of the L1 endonuclease domain from the N-terminal region of ORF2 does not affect reverse transcriptase activity as determined by this assay, and that approximately half of the ORF2 coding sequence from mouse A-type L1 elements is required for functional reverse transcriptase.
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PMID:Functional reverse transcriptases encoded by A-type mouse LINE-1: defining the minimal domain by deletion analysis. 966 81

New, noninvasive methods for the early detection of urothelial carcinomas of the urinary bladder are needed for the diagnosis, follow-up, and screening of patients with bladder cancer. Detection of the enzyme telomerase in urine could offer these new diagnostic possibilities. The standard technique for detecting telomerase activity is the telomeric repeat amplification protocol (TRAP assay). Because of the instability of the ribonucleoprotein telomerase in an aggressive medium, such as urine, investigations conducted to date have yielded nonuniform or even contradictory findings. This study compares the detection of human telomerase RNA (hTR) by reverse transcriptase-PCR (RT-PCR) with detection of telomerase activity by the TRAP assay in the diagnosis of urothelial carcinoma of the urinary bladder. Sedimented cells obtained from urine of 30 patients with urothelial carcinoma, 15 patients with benign urological disorders, 3 patients as part of follow-up for malignant disease, and 20 healthy subjects were examined for the presence of hTR and for telomerase activity (TRAP). In patients with bladder cancer, telomerase activity was detected by the TRAP assay in only 2 of 30 specimens (7%). However, increased levels of hTR were detected by RT-PCR in 25 of the same 30 cases (83%). For patients with benign urological disorders, such as urolithiasis or urinary tract infections, hTR was detected in samples obtained from 4 of 15 patients (27%). Low hTR expression levels were found in 15% of the healthy controls. The detection of hTR by RT-PCR represents a promising new method for detecting malignant cells in urine.
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PMID:Comparison of human telomerase RNA and telomerase activity in urine for diagnosis of bladder cancer. 971 24

The retrohoming of the yeast mtDNA intron aI1 occurs by a target DNA-primed reverse transcription (TPRT) mechanism in which the intron RNA reverse splices directly into the recipient DNA and is then copied by the intron-encoded reverse transcriptase. Here, we carried out biochemical characterization of the intron-encoded reverse transcriptase and site-specific DNA endonuclease activities required for this process. We show that the aI1 reverse transcriptase has high TPRT activity in the presence of appropriate DNA target sites, but differs from the closely related reverse transcriptase encoded by the yeast aI2 intron in being unable to use artificial substrates efficiently. Characterization of TPRT products shows that the fully reverse spliced intron RNA is an efficient template for cDNA synthesis, while reverse transcription of partially reverse spliced intron RNA is impeded by the branch point. Novel features of the aI1 reaction include a prominent open-circular product in which cDNAs are incorporated at a nick at the antisense-strand cleavage site. The aI1 endonuclease activity, which catalyzes the DNA cleavage and reverse splicing reactions, is associated with ribonucleoprotein particles containing the intron-encoded protein and the excised intron RNA. As shown for the aI2 endonuclease, both the RNA and protein components are used for DNA target site recognition, but the aI1 protein has less stringent nucleotide sequence requirements for the reverse splicing reaction. Finally, perhaps reflecting this relaxed target specificity, in vitro experiments show that aI1 can reverse splice directly into ectopic mtDNA transposition sites, consistent with the previously suggested possibility that this mechanism is used for ectopic transposition of group II introns in vivo.
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PMID:Group II intron mobility in yeast mitochondria: target DNA-primed reverse transcription activity of aI1 and reverse splicing into DNA transposition sites in vitro. 973 19

The reverse transcriptase telomerase is a ribonucleoprotein complex that adds telomeric repeats to chromosome ends, using a sequence within its endogenous RNA component as a template. Although templating domains of telomerase RNA have been studied in detail, little is known about the roles of the remaining residues, particularly in yeast. We examined the functions of nontemplate telomerase residues in the telomerase RNA of budding yeast Kluyveromyces lactis. Although approximately half of the RNA residues were dispensable for function, four specific regions were essential for telomerase action in vivo. We analyzed the effects of mutating these regions on in vivo function, in vitro telomerase activity, and telomerase RNP assembly. Deletion of two regions resulted in synthesis of stable RNAs that appeared unable to assemble into a stable RNP. Mutating a region near the 5' end of the RNA allowed RNP assembly but abolished enzymatic activity. Mutations in another specific small region of the RNA led to an inactive telomerase RNP with an altered RNA conformation.
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PMID:Specific telomerase RNA residues distant from the template are essential for telomerase function. 978 2

Telomerase is a ribonucleoprotein reverse transcriptase essential for the maintenance of telomere length. However, the available information concerning the biochemical and structural aspects of mammalian telomerases is scarce, primarily due to the low abundance of these enzymes and the difficulty and expense involved in its purification. To overcome these problems, we started to purify and characterize telomerase from bovine testis. Bovine telomerase was purified over columns of hydroxyapatite, DEAE-Sepharose, heparin-agarose, phenyl-agarose and spermine-agarose. In a sedimentation study performed using a 15-40% glycerol gradient, partially purified bovine telomerase exhibited an apparent molecular weight of more than 1000 kDa. One 435-bp RNA, bTR, was cloned from the most purified telomerase fraction and shown to be co-purified with telomerase activity in a glycerol gradient. bTR shares 83% similarity to the human telomerase RNA and 65% to the mouse telomerase RNA. A putative template region encompassing 10 nucleotides (5'-CUAACCCUAA-3') complementary to the mammalian telomere sequence (TTAGGG)n is located between nucleotides 38-47 of bTR. Besides, the bTR 5'-flanking region shares only three regulatory elements with that of hTR: a TATA-like sequence, a CCAAT box and an E1A-F box. Furthermore, the addition of in-vitro transcribed bTR reconstituted telomerase activity after removal of the endogenous bTR by micrococcal nuclease digestion. Northern blot analysis identified a single RNA of about 430-440 nucleotides expressed in the bovine testis and an immortalized bovine cell line. Taken together, these data strongly suggest that bTR is the RNA component of bovine telomerase.
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PMID:Molecular cloning of bovine telomerase RNA. 985 49

Simple sequence repeat telomeric DNA is maintained by a specialized reverse transcriptase, telomerase. The integral RNA subunit of telomerase contains a template region that determines the sequence added to chromosome ends. Aside from providing the template, little is known about the role of the telomerase RNA. In addition, no hypotheses have been suggested to account for the striking evolutionary divergence in size and sequence between telomerase RNAs of ciliates, yeasts, and mammals. We show that the two- to threefold increase in size of the mammalian telomerase RNAs relative to ciliate telomerase RNAs is due to the presence of an extra domain resembling a box H/ACA small nucleolar RNA (snoRNA). The human telomerase RNA (hTR) H/ACA domain is essential in vivo for hTR accumulation, hTR 3' end processing, and telomerase activity. By substituting the U64 box H/ACA snoRNA for the hTR H/ACA domain, we demonstrate that a heterologous snoRNA can function to promote chimeric RNA accumulation and 3' end processing but not telomerase activity. In addition, we show that maturation of full-length hTR and its assembly into active telomerase occur from an mRNA promoter-driven RNA polymerase II transcript but not from a U6 snRNA promoter-driven RNA polymerase III transcript. Finally, we show that a small percentage of hTR is associated with nucleoli. These results have implications for the biogenesis and structure of hTR and the human telomerase ribonucleoprotein complex. They also expand the structural and functional diversity of the box H/ACA snoRNA motif.
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PMID:A box H/ACA small nucleolar RNA-like domain at the human telomerase RNA 3' end. 985 80

In human immunodeficiency virus (HIV)-1 infection, decrease of telomere length is mainly found in CD8(+) T cells and not in CD4(+) T cells. Telomerase, a ribonucleoprotein enzyme that can synthesize telomeric sequence onto chromosomal ends, can compensate for telomere loss. Here, we investigated if telomerase activity could explain differential telomere loss of CD4(+) and CD8(+) T cells in HIV-1 infection. Telomerase activity was higher in CD8(+) than in CD4(+) T cells from HIV-infected patients, but still in the same range as in healthy controls, and upregulation after stimulation was comparable to normal. Telomerase activity in lymph node CD4(+) and CD8(+) T cells from HIV-infected patients was in the same range as that in CD4(+) and CD8(+) T cells from peripheral blood (PB) and was normal in unseparated bone marrow cells. Thus, our study did not provide evidence for compartmentalized elongation of telomeres in HIV infection. In patients treated with reverse transcriptase inhibitors, telomerase activity was inhibited, but this did not lead to accelerated loss of telomere length in vivo. Thus, differential telomere loss in CD4(+) and CD8(+) T cells in HIV-1 infection cannot be explained by telomerase activity.
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PMID:Normal T-cell telomerase activity and upregulation in human immunodeficiency virus-1 infection. 992 Aug 50

The MYC proto-oncogene encodes a ubiquitous transcription factor (c-MYC) involved in the control of cell proliferation and differentiation. Deregulated expression of c-MYC caused by gene amplification, retroviral insertion, or chromosomal translocation is associated with tumorigenesis. The function of c-MYC and its role in tumorigenesis are poorly understood because few c-MYC targets have been identified. Here we show that c-MYC has a direct role in induction of the activity of telomerase, the ribonucleoprotein complex expressed in proliferating and transformed cells, in which it preserves chromosome integrity by maintaining telomere length. c-MYC activates telomerase by inducing expression of its catalytic subunit, telomerase reverse transcriptase (TERT). Telomerase complex activity is dependent on TERT, a specialized type of reverse transcriptase. TERT and c-MYC are expressed in normal and transformed proliferating cells, downregulated in quiescent and terminally differentiated cells, and can both induce immortalization when constitutively expressed in transfected cells. Consistent with the recently reported association between MYC overexpression and induction of telomerase activity, we find here that the TERT promoter contains numerous c-MYC-binding sites that mediate TERT transcriptional activation. c-MYC-induced TERT expression is rapid and independent of cell proliferation and additional protein synthesis, consistent with direct transcriptional activation of TERT. Our results indicate that TERT is a target of c-MYC activity and identify a pathway linking cell proliferation and chromosome integrity in normal and neoplastic cells.
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PMID:Direct activation of TERT transcription by c-MYC. 998 78

Human telomerase is a ribonucleoprotein that maintains telomeres by adding TTAGGG tandem repeat sequences using RNA template of the enzyme. Telomerase activity is highly expressed in immortalized cells but not in most somatic cells, except for some renewal tissues, such as hematopoietic stem cells. Therefore, telomerase can be a target for anticancer drugs. Here we show that a fungus metabolite, alterperylenol, inhibits human telomerase activity. Alterperylenol inhibited telomerase activity (IC50 = 30 microM), but altertoxin I, a structurally related compound, did not affect it at 1 mM. Moreover, alterperylenol did not affect the activity of viral reverse transcriptase at 1 mM.
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PMID:Inhibition of human telomerase activity by alterperylenol. 1022 20


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