Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “mRNA”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 379 records · Page 21Linked to original sources
PubMed1999

[Effects of cyclosporin A combined with T4 on collagen IV production, coll alpha 1 (IV) mRNA and TGF beta 1 mRNA expression in cultured human mesangial cells].

OBJECTIVE: To investigate the effects of single cyclosporin A (CsA), tripcholorlide (T4) or both on renal fibrosis. METHODS: Collagen IV concentrations in the supernatants of human mesangial cells (HMCs) in culture were measured by ELISA. Collagen type IV alpha 1 mRNA (Coll alpha 1 IV mRNA) and transforming growth factor beta 1 (TGF beta 1) mRNA expression were tested by RT-PCR. RESULTS: CsA regulated HMC collagen IV production biphysically, i.e., 1.0 microgram/ml CsA increased collagen synthesis while 0.01-0.1 microgram/ml CsA decreased its formation. T4 enhanced collagen IV production in a dose dependent manner. Combination of the two drugs in lower doses could reach the same degree of immunosuppression as a higher dose of CsA did but with less fibrogenesis. Both CsA and T4 up-regulated HMC Coll alpha 1 (IV) mRNA and TGF beta 1 mRNA expression. The degree of Coll alpha 1 (IV) mRNA expression in different groups were in accordance with that of TGF beta 1 mRNA. CONCLUSIONS: Both CsA and T4 regulate extracellular matrix formation which may be mediated by TGF beta 1. Combination of the two drugs in lower dose results in less fibrosis than a higher dose of CsA.

Cells, Cultured↗
PubMed2002

[Correlation between the expression of mRNA of histones H2B and H4 and mRNA of kinin receptors B1 and B2 assessed by QRT-PCR in vulvar cancer].

OBJECTIVES: Histones are involved in process of proliferation and differentiation of the cells. In many lesions the expression of mRNA of histones is a marker of proliferation. The expression of kinin receptors B1 and B2 coexist with inflammatory processes, but in some studies the expression of these receptors was found to influence the growth and differentiation of different cells. DESIGN: In search of proliferation markers in squamous cell vulvar cancer we have analysed the correlation between the expression of the mRNA of histone H2B and H4 and mRNA of kinin receptors B1 and B2. MATERIALS AND METHODS: The specimen obtained from 46 women operated for squamous cell vulvar cancer stages I to IV according to FIGO were analysed. RNA was extracted and the number of mRNA copies were assessed by QRT-PCR using ABI PRISM 7700 (TaqMan). RESULTS: The results obtained in this study indicate a statistically significant correlation between mRNA B1 and mRNA H4 (p < 0.01), B2 and histone H4 (p < 0.01) B1 and histone H2B (p < 0.01). CONCLUSIONS: The expression of mRNA of histones and kinins receptors may reflects the dynamics of neoplastic growth in vulvar cancer.

Biomarkers, Tumor↗
PubMed2003

Evaluation of MDR1, LRP, MRP, and topoisomerase IIalpha gene mRNA transcripts before and after interferon-alpha, and correlation with the mRNA expression level of the telomerase subunits hTERT and TEP1 in five unselected human melanoma cell lines.

Intrinsic and acquired multidrug-resistance (MDR) and the activity of the enzyme telomerase have been demonstrated in human melanoma. A direct regulation of the MDR pathways and of telomerase by interpheron-alpha (IFN-alpha), which is currently used in the therapy of advanced cutaneous melanoma, has also been hypothesized. In this study, we used five melanoma cell lines not selected in vitro for drug resistance (Me665/2/21, Me665/2/60, HT-144, SK-MEL-28, and SK-MEL-5), which in a previous study, had shown different responses to IFN-alpha in terms of proliferation, apoptosis, telomerase activity and expression of mRNA for the human telomerase reverse transcriptase (hTERT). We investigated the expression of the multidrug resistance (MDR1) gene, multidrug resistance protein (MRP), lung resistance protein (LRP), topoisomerase IIalpha (Topo IIalpha), hTERT, and telomerase-associated protein (TEP1), which is shared by telomerase and vault MDR proteins at the mRNA expression level, using the reverse transcription-PCR (RT-PCR). All cell lines showed an intrinsic expression of hTERT, TEP1, and MDR gene transcripts (only MDR1 mRNA was under the detection level in SK-MEL-28 cells). After IFN-alpha exposure, we observed either no effect, a trend towards a decrease of hTERT, MRP, and Topo IIalpha, or an increase of TEP1, MDR1, and LRP mRNA expression in some cell lines. Effects were usually temporary and not always significant. No correlation was found between hTERT and TEP1 mRNA expression, whereas significant positive correlations were found between TEP1 and MDR1 mRNA, and between TEP1 and LRP mRNA. IFN-alpha modulates differently MDR gene transcripts in human melanoma cell lines. Positive correlation between TEP1 and LRP also seems to identify them as common targets of IFN-alpha effects.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2004

Increased expression of IGF II mRNA-binding protein 1 mRNA is associated with an advanced clinical stage and poor prognosis in patients with ovarian cancer.

The purpose of this study was to examine the expression of IGF II mRNA-binding protein (IMP-1, -2, and -3) mRNA in epithelial ovarian tumors, and to identify the association of IMP-1, -2, and -3 expression levels with patient survival. IMP mRNA expression levels were examined by semi-quantitative PCR in 59 epithelial ovarian tumors (8 adenomas, 5 LMP tumors, and 46 adenocarcinomas) and in 7 normal ovaries. Results of semiquantitative PCR were correlated with clinicopathological variables and overall survival. Human normal and tumor tissue cDNAs were included in all of the analyses. The IMP family mRNA expression was detected in almost all cancer tissues examined, including breast, lung, colon, prostatic, and ovarian carcinoma with the exception of pancreatic carcinoma. The mean value of the relative IMP-1 mRNA expression ratio was significantly higher in both ovarian cancer and adenoma samples compared to normal ovarian samples (p<0.05). IMP-2 and IMP-3 expression levels were significantly higher in both ovarian cancer and ovarian LMP tumor samples compared to either ovarian adenomas or to normal ovary samples (p<0.05). A significantly higher IMP-1 mRNA expression level was observed in patients with an advanced clinical stage (p=0.015) and high histological grade (p=0.023). Log-rank testing showed that IMP-1 overexpression (p=0.0398) and an advanced clinical stage (p=0.0050) were significantly correlated with poor patient survival, whereas neither IMP-2 nor IMP-3 overexpression were associated with poor prognoses. In multivariate analysis, IMP-1 overexpression lost its significance, whereas the clinical stage (p=0.0432) remained significantly associated with overall survival. IMP mRNA expression levels might play an important role in ovarian cancer development and progression, and IMP-1 overexpression is a prognostic marker for patients with ovarian cancer.

DNA, Complementary↗
PubMed2005

[The effect of ribozyme specially cleaving per1 mRNA on c-fos mRNA and its expression in hippocampus of morphine addicted mice].

OBJECTIVE: To study the change of c-fos mRNA and protein in hippocampus of morphine addicted mice after injected with ribozyme specially cleaving per1 mRNA. METHOD: The recombined plasmid pcDNA 3.1-per1RZ DNA was injected into the ventricles of morphine addicted mice to transcript the corresponding ribozyme which cleaves per1 mRNA particularly. And then, the brains of mice were fixed by perfusion. The level of c-fos mRNA was assayed by in situ hybridization and c-fos protein was detected by immunohistochemical staining. RESULT: The level of c-fos mRNA and protein decreased after injection of the recombined plasmid pcDNA 3.1-per1RZ DNA expressing the ribozyme cleaving per1 mRNA. CONCLUSION: The ribozyme specially cleaving per1 mRNA has potential function in inhibiting the transcription and expression of c-fos and blocking the morphine addiction.

Animals↗
PubMed1991

Decreased catalytic subunit mRNA levels and altered catalytic subunit mRNA structure in a cAMP-resistant Chinese hamster ovary cell line.

The mechanisms responsible for decreased levels of cAMP-dependent protein kinase activity in a mutant Chinese hamster ovary cell line have been examined. The cAMP-resistant Chinese hamster ovary 10260 cell line was found to possess only 20% of the cAMP-dependent protein kinase activity found in wild-type cells. The presence of decreased concentrations of the catalytic subunit in these cells was confirmed through binding studies using a radiolabeled, heat-stable inhibitor of the kinase. Cloned Chinese hamster ovary catalytic subunit cDNAs were isolated, characterized, and used as hybridization probes to examine the relative concentrations of catalytic subunit mRNAs in the wild-type and 10260 cell lines. A 40-50% decrease in the concentration of the mRNA for the C alpha isozyme of the catalytic subunit was observed in 10260 cells, as compared with wild-type. This decrease in catalytic subunit mRNA concentration probably accounts for a portion of the decreased kinase activity in the mutant cells. Further analysis of C alpha mRNA by polymerase chain reaction confirmed the decreased expression of C alpha mRNA in 10260 cells and further demonstrated the presence of two different species of C alpha mRNA in the 10260 cells. One species of C alpha cDNAs was indistinguishable from the wild-type cDNA, but the other species was shorter. Nucleotide sequence analysis of the amplified cDNAs led to the identification of a 191-base pair deletion in the shorter cDNA. Gene transfer studies using wild-type and 10260 C alpha cDNAs demonstrated that the longer cDNA from the 10260 cells produced wild-type activity, but the shorter cDNA was inactive. These studies suggest that at least two alterations in gene expression are responsible for decreased cAMP-dependent protein kinase activity in the 10260 cell line. One alteration results in an approximately 2-fold decrease in the concentrations of C alpha mRNA in the cells. The other change produces two species of C alpha mRNA; one of the C alpha mRNAs does not encode an active kinase.

Amino Acid Sequence↗
PubMed1991

mRNA degradation by processive 3'-5' exoribonucleases in vitro and the implications for prokaryotic mRNA decay in vivo.

Two 3'-5' exoribonucleases, polynucleotide phosphorylase and ribonuclease II play a central role in the degradation of bacterial mRNA to ribonucleotides. Sequences with the potential to form stem-loop structures can stabilize upstream mRNA against 3'-5' exoribonucleolytic attack in vivo by blocking the processive activities of these enzymes. For many mRNA species stem-loop structures appear to provide a very efficient block to decay from the 3' end, such that the rate-determining step for mRNA decay occurs elsewhere in the transcript. We have examined the stalling of 3'-5' exoribonucleases at stem-loop structures in vitro. Although stem-loop structures alone can impede the progress of both enzymes, the duration of stalling at these structures in vitro is insufficient to account for the increased half-lives that they confer on mRNA in vivo. These data suggest that an additional factor, such as a stem-loop binding protein, is required for stabilization of mRNA by stem-loop structures in vivo. The implications for the regulation of mRNA stability are discussed.

Amino Acid Sequence↗
PubMed1984

Cloning of cDNA sequences for murine ATP-citrate lyase. Construction of recombinant plasmids using an immunopurified mRNA template and evidence for the nutritional regulation of ATP-citrate lyase mRNA content in mouse liver.

Mouse liver mRNA that was enriched in sequences coding for ATP-citrate lyase by polysome immunoadsorption was used as a template for cDNA synthesis. Double-stranded cDNA sequences were inserted into the plasmid pBR322 and cloned in Escherichia coli RR1. Twenty-seven plasmids containing putative cDNA sequences for ATP-citrate lyase were identified by differential hybridization with single-stranded 32P-cDNAs synthesized from immunopurified mRNA, sucrose gradient-purified ATP-citrate lyase mRNA, and mRNA isolated from the livers of mice that were nutritionally induced or de-induced for ATP-citrate lyase biosynthesis. A subgroup of five recombinant plasmids was characterized further in hybridization-selection experiments. Each of these plasmids positively selected ATP-citrate lyase mRNA as determined by in vitro translation and specific immunoprecipitation. The length of ATP-citrate lyase mRNA was estimated to be 4900 bases in a Northern blot analysis. A 32P-cDNA probe derived from a 1500-base pair insert was used to investigate the basis for the 20-30-fold induction of ATP-citrate lyase that occurs when starved animals are fed a high carbohydrate/low fat diet. Dot-blot hybridization analysis disclosed that the relative content of liver ATP-citrate lyase mRNA increased 25-fold after 15 h of refeeding, indicating that the synthesis of the lipogenic enzyme is controlled at a pretranslational level in the nutritional paradigm.

ATP Citrate (pro-S)-Lyase↗
PubMed1983

mRNA secondary structure as a determinant in cap recognition and initiation complex formation. ATP-Mg2+ independent cross-linking of cap binding proteins to m7I-capped inosine-substituted reovirus mRNA.

Polypeptides of Mr = 50,000 and 80,000 in rabbit reticulocyte initiation factor preparations can be specifically cross-linked to the oxidized t' cap structure of native reovirus mRNA in an ATP-Mg2+-dependent manner (Sonenberg, N., Guertin, D., Cleveland, D., and Trachsel, H. (1981) Cell 27, 563-572). However, specific cross-linking of these polypeptides can occur in the absence of ATP-Mg2+ when m7I-capped inosine substituted mRNA, which contains less secondary structure than native reovirus mRNA, is used. We also found, using wheat germ extract, that inhibition of initiation complex formation by high salt concentrations is directly related to the degree of secondary structure of the mRNA. Binding of ribosomes to bromouridine-substituted reovirus mRNA is severely inhibited at high K+ concentrations, while binding to inosine-substituted mRNA is only slightly inhibited and binding of native reovirus mRNA is inhibited to an intermediate degree. The results are consistent with the hypothesis that cap recognition factors mediate an ATP-dependent melting of secondary structures involving 5' proximal sequences to the initiation codon in order to facilitate binding of ribosomes during translation initiation.

Adenosine Triphosphate↗
PubMed1984

Analysis of mRNA coding for blood-stage antigens of a rodent malarial parasite, Plasmodium yoelii: mRNA coding for a possible protective antigen purify as poly A-.

We have analyzed mRNA coding for blood-stage antigens of Plasmodium yoelii by using cellfree translation of poly A+ and poly A- RNA in conjunction with immunoprecipitations. Most of the antigens recognized by mouse hyperimmune serum to P. yoelii were coded by poly A+ mRNA ranging in size from 15S to 28S. However, certain P. yoelii antigens, notably those with m.w. greater than 150 kilodaltons (kd), were coded by mRNA that purified as being poly A-. Antigens recognized by a protective monoclonal antibody (McAb) were coded by such operationally poly A- RNA. Three polypeptides apparently coded by different poly A- RNA were immunoprecipitated by this McAb. With the use of another McAb, a poly A+ mRNA of about 19S was identified as coding for a polypeptide of 46 kd synthesized in cellfree translation reactions. The same McAb recognized a 34 kd polypeptide in metabolically labeled polypeptides of P. yoelii. This antigen appeared to be processed in vivo but not in vitro. The observation that some mRNA of P. yoelii purify as being poly A- has significant implications for the construction of cDNA libraries that employ poly A+ mRNA of malarial parasites: if it applies to other species of plasmodia, some potentially important operationally poly A- mRNA may not be represented in such libraries.

Animals↗
PubMed1984

Beta O-39 thalassemia gene: a premature termination codon causes beta-mRNA deficiency without affecting cytoplasmic beta-mRNA stability.

The function of a beta O-thalassemia globin gene with a premature termination codon at position 39 was studied in tissue culture cells using plasmid expression vectors. The thalassemic globin gene was isolated by molecular cloning from the bone marrow DNA of an Italian patient with severe thalassemia. Sequences upstream of the normal beta-globin gene and the beta O-39 globin gene were removed to 127 nucleotides (nt) 5' to the start site of transcription, thereby eliminating uncharacterized DNA sequences but preserving promoter function. To create a hybrid gene differing from the normal by only the single nt change in codon 39, the truncated 5' end of the beta O-39 gene was recombined with the 3' end of the normal gene. The beta O-39 substitution resulted in a 6-14-fold reduction in cytoplasmic RNA accumulation in transfected monkey kidney or HeLa cells. This decrease in mRNA did not appear to be due to instability, as the beta-mRNA present 24-36 hours after transfection was stable for up to 24 hours in the presence of actinomycin D. In the presence of actinomycin D, the globin mRNA precursor disappeared, suggesting that globin gene transcription had been effectively blocked. We also examined a second globin mRNA with a premature termination codon; this RNA that arises from incorrect splicing of the transcript of a beta + -24 thalassemia gene was as stable as the correctly spliced species. Thus, the presence of a premature termination codon does not necessarily alter cytoplasmic mRNA stability, nor does cytoplasmic instability account for the quantitative deficiency of beta-globin mRNA. Our observations suggest that a premature termination codon alters intranuclear stability and/or nuclear to cytoplasmic transport of the beta-globin mRNA.

Codon↗
PubMed1995

Localization of novel corticotropin-releasing factor receptor (CRF2) mRNA expression to specific subcortical nuclei in rat brain: comparison with CRF1 receptor mRNA expression.

Corticotropin-releasing factor (CRF) is the primary factor involved in controlling the release of ACTH from the anterior pituitary and also acts as a neurotransmitter in a variety of brain systems. The actions of CRF are mediated by G-protein coupled membrane bound receptors and a high affinity CRF receptor, CRF1, has been previously cloned and functionally characterized. We have recently isolated a cDNA encoding a second member of the CRF receptor family, designated CRF2, which displays approximately 70% homology at the nucleotide level to the CRF1 receptor and exhibits a distinctive pharmacological profile. The present study utilized in situ hybridization histochemistry to localize the distribution of CRF2 receptor mRNA in rat brain and pituitary gland and compared this with the distribution of CRF1, receptor expression. While CRF1 receptor expression was very high in neocortical, cerebellar, and sensory relay structures, CRF2 receptor expression was generally confined to subcortical structures. The highest levels of CRF2 receptor mRNA in brain were evident within the lateral septal nucleus, the ventromedial hypothalamic nucleus and the choroid plexus. Moderate levels of CRF2 receptor expression were evident in the olfactory bulb, amygdaloid nuclei, the paraventricular and suraoptic nuclei of the hypothalamus, the inferior colliculus and 5-HT-associated raphe nuclei of the midbrain. CRF2-expressing cells were also evident in the bed nucleus of the stria terminalis, the hippocampal formation and anterior and lateral hypothalmic areas. In addition, CRF2 receptor mRNA was also found in cerebral arterioles throughout the brain. Within the pituitary gland, CRF2 receptor mRNA was detectable only at very low levels in scattered cells while CRF1 receptor mRNA was readily detectable in anterior and intermediate lobes. This heterogeneous distribution of CRF1 and CRF2 receptor mRNA suggests distinctive functional roles for each receptor in CRF-related systems. The CRF1 receptor may be regarded as the primary neuroendocrine pituitary CRF receptor and important in cortical, cerebellar and sensory roles of CRF. The anatomical distribution of CRF2 receptor mRNA indicates a role for this novel receptor in hypothalamic neuroendocrine, autonomic and general behavioral actions of central CRF.

Animals↗
PubMed1998

mRNA stabilization by the ompA 5' untranslated region: two protective elements hinder distinct pathways for mRNA degradation.

The 5' untranslated region (UTR) of the long-lived Escherichia coli ompA transcript functions as an mRNA stabilizer that can prolong the cytoplasmic lifetimes of a variety of messages to which it is fused. Previous studies have identified two domains of this 5' UTR that together are responsible for its stabilizing effect. One is a 5'-terminal stem-loop. The other is a single-stranded RNA segment (ss2) that contains a ribosome binding site highly complementary to 16S ribosomal RNA. Here we report a detailed investigation of the function of these two stabilizing elements. Our data indicate that mRNA protection by a 5' stem-loop requires no sequence features or thermodynamic stability beyond the minimum necessary for stem-loop formation. Stabilization by ss2 appears to result not from a high frequency of translation initiation, but rather from a high degree of occupancy of this 5' UTR segment by bound ribosomes. Although close spacing of translating ribosomes is not critical for message stabilization by the ompA 5' UTR, mRNA longevity does require the periodic passage of ribosomes through the protein-coding region. Unlike bound ribosomes, which hinder mRNA cleavage by RNase E, the 5' stem-loop appears to impede degradation of ompA mRNA via a distinct pathway that is RNase E-independent. These findings imply that the ompA 5' UTR prolongs mRNA longevity by impeding multiple pathways for mRNA degradation.

Bacterial Outer Membrane Proteins↗
PubMed1996

The protein that binds the 3' end of histone mRNA: a novel RNA-binding protein required for histone pre-mRNA processing.

Replication-dependent histone mRNAs are not polyadenylated but end in a conserved 26-nucleotide structure that contains a stem-loop. Much of the cell cycle regulation of histone mRNA is post-transcriptional and is mediated by the 3' end of histone mRNA. The stem-loop binding protein (SLBP) that binds the 3' end of histone mRNA is a candidate for the factor that participates in most, if not all, of the post-transcriptional regulatory events. We have cloned the cDNA for the SLBP from humans, mice, and frogs, using the recently developed yeast three-hybrid system. The human SLBP is a 31-kD protein and contains a novel RNA-binding domain, which has been mapped to a 73-amino-acid region of the protein. The cloned SLBP is the protein bound to the 3' end of histone mRNA as antibodies specific for the SLBP remove all specific binding activity from nuclear and polyribosomal extracts. These depleted extracts do not cleave histone pre-mRNA efficiently, demonstrating that the SLBP is required for efficient histone pre-mRNA processing.

Amino Acid Sequence↗
PubMed2002

Deletion of individual mRNA capping genes is unexpectedly not lethal to Candida albicans and results in modified mRNA cap structures.

Eukaryotic mRNAs are modified at the 5' end with a cap structure. In fungal cells, the formation of the mRNA cap structure is catalyzed by three enzymes: triphosphatase, guanylyltransferase, and methyltransferase. Fungal capping enzymes have been proposed to be good antifungal targets because they differ significantly from their human counterparts and the genes encoding these enzymes are essential in Saccharomyces cerevisiae. In the present study, Candida albicans null mutants were constructed for both the mRNA triphosphatase-encoding gene (CET1) and the mRNA methyltransferase encoding gene (CCM1), proving that these genes are not essential in C. albicans. Heterozygous deletions were generated, but no null mutants were isolated for the guanylyltransferase-encoding gene (CGT1), indicating that this gene probably is essential in C. albicans. Whereas these results indicate that Cet1p and Ccm1p are not ideal molecular targets for development of anticandidal drugs, they do raise questions about the capping of mRNA and translation initiation in this fungus. Southern blot analysis of genomic DNA indicates that there are not redundant genes for CET1 and CCM1 and analysis of mRNA cap structures indicate there are not alternative pathways compensating for the function of CET1 or CCM1 in the null mutants. Instead, it appears that C. albicans can survive with modified mRNA cap structures.

Acid Anhydride Hydrolases↗
PubMed2004

The oligo(A) tail on histone mRNA plays an active role in translational silencing of histone mRNA during Xenopus oogenesis.

Metazoan replication-dependent histone mRNAs end in a stem-loop sequence. The one known exception is the histone mRNA in amphibian oocytes, which has a short oligo(A) tail attached to the stem-loop sequence. Amphibian oocytes also contain two proteins that bind the 3' end of histone mRNA: xSLBP1, the homologue of the mammalian SLBP, and xSLBP2, which is present only in oocytes. xSLBP2 is an inhibitor of histone mRNA translation, while xSLBP1 activates translation. The short A tail on histone mRNAs appears at stage II to III of oogenesis and is present on histone mRNAs throughout the rest of oogenesis. At oocyte maturation, the oligo(A) tail is removed and the xSLBP2 is degraded, resulting in the activation of translation of histone mRNA. Both SLBPs bind to the stem-loop with the oligo(A) tail with similar affinities. Reporter mRNAs ending in the stem-loop with or without the oligo(A) tail are translated equally well in a reticulocyte lysate, and their translation is stimulated by the presence of xSLBP1. In contrast, translation of the reporter mRNA with an oligo(A) tail is not activated in frog oocytes in response to the presence of xSLBP1. These results suggest that the oligo(A) tail is an active part of the translation repression mechanism that silences histone mRNA during oogenesis and that its removal is part of the mechanism that activates translation.

Adenine Nucleotides↗
PubMed1992

mRNA capping enzyme. Isolation and characterization of the gene encoding mRNA guanylytransferase subunit from Saccharomyces cerevisiae.

The highly purified yeast mRNA capping enzyme is composed of two separate chains of 52 (alpha) and 80 kDa (beta), responsible for the activities of mRNA guanylyltransferase and RNA 5'-triphosphatase, respectively (Itoh, N., Yamada, H., Kaziro, Y., and Mizumoto, K. (1987) J. Biol. Chem. 262, 1989-1995). The gene encoding the mRNA guanylyltransferase subunit (alpha subunit), CEG1, has been isolated by immunological screening of a yeast genomic expression library in lambda gt11 with polyclonal antibodies directed against purified yeast capping enzyme. The identity of CEG1 was confirmed by epitope selection and by expressing the gene in Escherichia coli to give a catalytically active mRNA guanylyltransferase. The gene is present in one copy per haploid genome, and encodes a polypeptide of 459 amino acid residues. From its primary structure as well as its mRNA size, it was concluded that the alpha and the beta subunits of yeast mRNA capping enzyme are encoded by two separate genes, not as a fused protein. CEG1 is located on the chromosome VII by a pulse-field gel electrophoresis. Gene disruption experiment indicated that CEG1 is essential for the growth of yeast. We have also found another open reading frame (ORF2) which lies in close proximity to CEG1 in our clones and encodes a 450 amino acid-polypeptide of yet unknown function.

Amino Acid Sequence↗