Removal of double-stranded contaminants from RNA transcripts: synthesis of adenovirus VA RNA<sub>1</sub>from a T7 vector

Mellits, Kenneth H.; Pe’ery, Tsafi; Manche, Lisa; Robertson, Hugh D.; Mathews, Michael B.

doi:10.1093/nar/18.18.5401

Cited by 61 publications

(71 citation statements)

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“…Kinase Assays-Reactions (20 l) containing 2.5 Ci of [␥-32 P]ATP (ICN Biomedical Inc., Costa Mesa, CA) and 0.5 l of PKR (approximately 5 ng) purified to the mono-S stage (46) were conducted as described previously (47) in the presence of dsRNA derived from reovirus (a gift from A. Shatkin). Kinase reactions (20 l total volume) containing PKC (10 ng) were carried out as described by the manufacturer (Upstate Biotechnology Inc., Lake Placid, NY).…”

Section: Methodsmentioning

confidence: 99%

The Tat Protein of Human Immunodeficiency Virus Type 1 Is a Substrate and Inhibitor of the Interferon-induced, Virally Activated Protein Kinase, PKR

Brand

Kobayashi

Mathews

1997

Journal of Biological Chemistry

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135

102

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Section: Methodsmentioning

confidence: 99%

The Tat Protein of Human Immunodeficiency Virus Type 1 Is a Substrate and Inhibitor of the Interferon-induced, Virally Activated Protein Kinase, PKR

Brand

Kobayashi

Mathews

1997

Journal of Biological Chemistry

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135

102

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“…VA RNA, is a short (160 nucleotide) RNA which is synthesized by RNA polymerase HI. It binds to DAI, but unlike dsRNA it does not activate the enzyme (2,3,(17)(18)(19)(20).…”

Section: Introductionmentioning

confidence: 99%

Structural features of adenovirus 2 virus-associated RNA required for binding to the protein kinase DAI

Clarke

Pe’ery

et al. 1994

Nucl Acids Res

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“…Longer dsRNAs (354 bp) were synthesized by transcription of the pGEM.GC plasmid (42 (29) were conducted essentially as described by Mellits et al (42). The enzyme was added last to the other reaction components assembled on ice.…”

mentioning

confidence: 99%

Interactions between double-stranded RNA regulators and the protein kinase DAI.

Manche¹,

Green²,

Schmedt³

et al. 1992

Mol. Cell. Biol.

461

402

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The interferon-induced protein kinase DAI, the double-stranded RNA (dsRNA)-activated inhibitor of translation, plays a key role in regulating protein synthesis in higher cells. Once activated, in a process that involves autophosphorylation, it phosphorylates the initiation factor eIF-2, leading to inhibition of polypeptide chain initiation. The In mammalian cells, a regulatory mechanism involving an RNA-activated protein kinase and the eukaryotic initiation factor 2 (eIF-2) has been intensively studied. This initiation factor forms a ternary complex with GTP and Met-tRNAF and delivers the initiator tRNA to the ribosomal site of protein synthesis initiation. Discharged eIF-2 is subsequently released as a complex with GDP which must be replaced with GTP to permit the formation of another ternary complex in preparation for a further round of initiation. The factor is composed of three dissimilar subunits, cx, I, and -y. Phosphorylation of a single residue, serine-51 of the a subunit, inhibits translation by trapping a second initiation factor, the guanosine nucleotide exchange factor (or eIF-2B), which is required to catalyze the substitution of GTP for GDP in the discharged eIF-2 complex. Phosphorylation of sufficient eIF-2 can sequester all of the guanosine nucleotide exchange factor, thereby preventing eIF-2 recycling and halting the initiation pathway.In mammals, two protein kinases are capable of phosphorylating the a subunit of eIF-2 in this way (reviewed in references 20, 37, and 46). One of them, the heme-controlled repressor, is found chiefly in reticulocytes. It is activated by the absence of hemin, as well as by other stimuli, and serves to prevent the accumulation of globin in the absence of iron or heme. A second kinase, the double-stranded RNA-activated inhibitor (DAI; also referred to as P1 kinase, p68 kinase, P1/eIF-2a kinase, and PKdS, etc.) is present in a wide range of tissues. DAI is an important element in the host antiviral response, and its synthesis is induced at the transcriptional level by interferon (reviewed in references 21, 54, 56, and 59). The enzyme is ribosome associated (11,34) (8,24,26,36,47,60). It has also been implicated in cellular differentiation (23,52), in the inhibition of cell proliferation (6, 51), in the heat shock response (10), and possibly in transcriptional induction (61, 64). Moreover, in yeast cells, the related protein kinase GCN2 mediates the growth response to amino acid starvation (9). As its name implies, DAI is activated by doublestranded RNA (dsRNA). Other polyanions such as heparin can also activate it, while small, highly structured RNA molecules such as adenovirus VA RNA suppress its activation (38). Thus, DAI is a pivotal cellular regulatory enzyme whose level and activity are modulated by factors of both viral and cellular origin.The interactions between DAI and its RNA effectors are complicated and incompletely understood. The kinase is activated by dsRNA but not by DNA or DNA-RNA hybrids (22,32,35,58). Single-stranded RNA, either synthetic or natur...

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Removal of double-stranded contaminants from RNA transcripts: synthesis of adenovirus VA RNA₁from a T7 vector

Cited by 61 publications

References 50 publications

The Tat Protein of Human Immunodeficiency Virus Type 1 Is a Substrate and Inhibitor of the Interferon-induced, Virally Activated Protein Kinase, PKR

The Tat Protein of Human Immunodeficiency Virus Type 1 Is a Substrate and Inhibitor of the Interferon-induced, Virally Activated Protein Kinase, PKR

Structural features of adenovirus 2 virus-associated RNA required for binding to the protein kinase DAI

Interactions between double-stranded RNA regulators and the protein kinase DAI.

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Removal of double-stranded contaminants from RNA transcripts: synthesis of adenovirus VA RNA1from a T7 vector

Cited by 61 publications

References 50 publications

The Tat Protein of Human Immunodeficiency Virus Type 1 Is a Substrate and Inhibitor of the Interferon-induced, Virally Activated Protein Kinase, PKR

The Tat Protein of Human Immunodeficiency Virus Type 1 Is a Substrate and Inhibitor of the Interferon-induced, Virally Activated Protein Kinase, PKR

Structural features of adenovirus 2 virus-associated RNA required for binding to the protein kinase DAI

Interactions between double-stranded RNA regulators and the protein kinase DAI.

Contact Info

Product

Resources

About

Removal of double-stranded contaminants from RNA transcripts: synthesis of adenovirus VA RNA₁from a T7 vector