Statistical Analysis of the 5′ Untranslated Region of Human mRNA Using “Oligo-Capped” cDNA Libraries

Suzuki, Yutaka; Ishihara, Daisuke; Sasaki, Masahide; Nakagawa, H; Hata, Hiroko; Tsunoda, Takeshi; Watanabe, Masao; Komatsu, Takami; Ota, Toshio; Isogai, Takao; Suyama, Akira; Sugano, Sumio

doi:10.1006/geno.2000.6076

Cited by 134 publications

(144 citation statements)

References 40 publications

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“…The cDNA clone mncb-0146 (GeneBank Accession #: AU035162) was from the Sugano mouse brain mncb library (Suzuki et al, 2000). Double-strand DNA sequencing showed that this clone encodes the full-length mouse PTEN protein and confirmed that the 5 0 -UTR sequence (À828 to À1) was identical to that shown in GeneBank (Accession # NM_008960).…”

Section: Construction Of Plasmidsmentioning

confidence: 79%

Regulation of constitutive expression of mouse PTEN by the 5′-untranslated region

et al. 2003

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PTEN tumor suppressor serves as a major negative regulator of survival signaling mediated by PI3 kinase/ AKT/protein kinase B pathway, and is inactivated in various human tumors. Elucidation of mechanisms responsible for PTEN expression is important for providing insight into strategies to control the loss of PTEN expression in human cancers. Although recent studies suggested that p53 and Egr-1 can modulate induced PTEN expression, the mechanism responsible for ubiquitous constitutive expression of PTEN remains elusive. PTEN mRNA contains a highly conserved and GC-rich 5 0 -untranslated region (5 0 -UTR). Recently, it has been shown that the long 5 0 -UTR sequences of several growthregulated mRNAs contain promoters that can generate mRNAs with shorter 5 0 -UTRs. In this paper, we tested whether the 5 0 -UTR sequence of mouse PTEN contains a promoter that is responsible for constitutive expression of PTEN. We found that the long 5 0 -UTR sequence of mouse PTEN severely inhibits translation of PTEN and a heterologous gene firefly luciferase. Deletion of the most 5 0 -UTR sequence would enhance translation efficiency 100-fold. We also showed that the 5 0 -UTR sequence of mouse PTEN does not have an internal ribosome entry site (IRES) that can mediate cap-independent initiation of translation. Instead, we found that the 5 0 -UTR sequence of mouse PTEN contains a strong promoter that drives the production of a transcript with shorter 5 0 -UTRs, which can be translated with higher efficiency. This promoter was mapped to the region between À551 and À220 bases upstream of the translation start codon. Cotransfection analysis using Drosophila SL2 cells showed that Sp1 is one of the major transcription factors that can constitutively activate this promoter. Two endogenous PTEN transcripts with 5 0 -UTRs of 193 and 109 bases were found in DU145 and H226 cell lines. Based on these observations, we conclude that the PTEN expression may be regulated at both transcriptional and translational levels, and that the 5 0 -UTR sequence of PTEN contains a promoter that is responsible for constitutive PTEN expression.

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Section: Construction Of Plasmidsmentioning

confidence: 79%

Regulation of constitutive expression of mouse PTEN by the 5′-untranslated region

et al. 2003

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“…Although the sequence databases do not contain good information on the 5 ¶ ends of most genes, compilations of genes with known 5 ¶ UTRs indicate that these elements are relatively common. It has been estimated that 10% to 30% of genes contain one or more uORFs and 15% to 53% of mRNAs contain upstream start codons (102,103). Thus, a large number of transcripts can be potentially influenced through this type of mechanism.…”

Section: Selective Gene Expression Mediated By Eif2 Regulation Duringmentioning

confidence: 99%

“Translating” Tumor Hypoxia: Unfolded Protein Response (UPR)–Dependent and UPR-Independent Pathways

Koumenis

Wouters

2006

Molecular Cancer Research

205

170

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Poor oxygenation (hypoxia) is present in the majority of human tumors and is associated with poor prognosis due to the protection it affords to radiotherapy and chemotherapy. Hypoxia also elicits multiple cellular response pathways that alter gene expression and affect tumor progression, including two recently identified separate pathways that strongly suppress the rates of mRNA translation during hypoxia. The first pathway is activated extremely rapidly and is mediated by phosphorylation and inhibition of the eukaryotic initiation factor 2A. Phosphorylation of this factor occurs as part of a coordinated endoplasmic reticulum stress response program known as the unfolded protein response and activation of this program is required for hypoxic cell survival and tumor growth. Translation during hypoxia is also inhibited through the inactivation of a second eukaryotic initiation complex, eukaryotic initiation factor 4F. At least part of this inhibition is mediated through a Redd1 and tuberous sclerosis complex 1/2 -dependent inhibition of the mammalian target of rapamycin kinase. Inhibition of mRNA translation is hypothesized to affect the cellular tolerance to hypoxia in part by promoting energy homeostasis. However, regulation of translation also results in a specific increase in the synthesis of a subset of hypoxia-induced proteins. Consequently, both arms of translational control during hypoxia influence gene expression and phenotype. These hypoxic response pathways show differential activation requirements that are dependent on the level of oxygenation and duration of hypoxia and are themselves highly dynamic. Thus, the severity and duration of hypoxia can lead to different biological and therapeutic consequences. (Mol Cancer Res 2006;4(7):423 -36)

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“…However, the CEP4 nucleotide sequence in the database was incomplete in the untranslated region; therefore, we searched for a full-length Borg4 cDNA clone in a full-length enriched cDNA library from human adult ileum mucosa (Suzuki et al 2000) and found the clone KAIA1777. The insert length of KAIA1777 was 2732 bp and the coding sequence shared 80% identity with that of mouse Borg4.…”

Section: Resultsmentioning

confidence: 99%

“…The mouse Borg4 cDNA clone (MNCb-0220; DDBJ/ EMBL/GenBank accession number AB035088; the 5Ј-part of the sequence was deposited as accession number AU035226) and the putative human orthologue clone (KAIA1777; accession number AB042237) were derived from a mouse brain and a human adult ileum mucosa cDNA library made by the oligo-capping method (Suzuki et al 1997;Suzuki et al 2000). The method allowed us to replace the cap structure of mRNA with synthetic oligonucleotides (5Ј-oligo), which could then serve as the sequence tag for the mRNA start site.…”

Section: Dna Sequencingmentioning

confidence: 99%

Sequence analysis, gene expression, and chromosomal assignment of mouse Borg4 gene and its human orthologue

et al. 2000

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The Borg (binder of Rho GTPases) family proteins interact with CDC42 and TC10 in a guanosine triphosphate (GTP)-dependent manner. We have isolated a full-length cDNA of the mouse Borg4 gene, which is a member of this family. Sequence analysis revealed that this gene encoded a putative 349-amino acid protein. By reverse transcription -coupled polymerase chain reaction (RT-PCR) analysis, we observed that Borg4 was expressed ubiquitously in adult tissues. Additionally, we determined the entire cDNA sequence of the putative human Borg4 orthologue. By fluorescence in situ hybridization, mouse Borg4 and the putative human orthologue have been assigned to mouse chromosome 11E and human chromosome 17q24-25, which has been described as syntenic to the mouse region.

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Statistical Analysis of the 5′ Untranslated Region of Human mRNA Using “Oligo-Capped” cDNA Libraries

Cited by 134 publications

References 40 publications

Regulation of constitutive expression of mouse PTEN by the 5′-untranslated region

Regulation of constitutive expression of mouse PTEN by the 5′-untranslated region

“Translating” Tumor Hypoxia: Unfolded Protein Response (UPR)–Dependent and UPR-Independent Pathways

Sequence analysis, gene expression, and chromosomal assignment of mouse Borg4 gene and its human orthologue

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