Nascent RNA transcripts

DNA stores genetic information in a stable form that can be readily replicated. However, the expression of this genetic information requires its How from DNA to RNA to protein, as was introduced in Chapter 4, The present chapter deals with how RNA is synthesized and then modified to prepare for its translation into protein. We begin with transcription in prokaryotes and focus on the three stages of transcription promoter binding and initiation, elongation of the nascent RNA transcript, and termination at the end of the gene. 

Portion of a "lampbrush chromosome" from an oocyte of the newt Nophthalmus viridescens] hnRNP protein associated with nascent RNA transcripts fluoresces red after staining with a monoclonal antibody. [Courtesy... 

Nascent RNA transcripts from protein-coding genes and mRNA processing Intermediates, collectively referred to as pre-mRNA, do not exist as free RNA molecules in the nuclei of eukaryotic cells. From the time nascent transcripts first emerge from RNA polymerase II until mature mRNAs are transported into the cytoplasm, the RNA molecules are associated with an abundant set of nuclear proteins. These... 

RNA editing is a post-transcriptional event that changes one canonical nucleotide into another in nascent RNA transcripts, especially in mRNAs. Since this has the potential to change the amino acid identity of the pertinent codon, RNA editing has been shown to play an extremely important role in generating protein diversity and/or regulating translation specificity. Of all the RNA editing events, deamination is probably the one that has been most extensively studied. In this section, two representative deamination modifications, adenosine to inosine (A to I) and cytidine to uridine (C to U) conversions are discussed. 

Huang, J. and van der Ploeg, L. H. T. (1991) Maturation of polycistronic pre-mRNA in Trypanosoma brucei analysis of trans-splicing and poly (A) addition at nascent RNA transcripts from the hsp 70 locus. Mol. Cell Biol. 11 3180-3190. 

The basic enzymatic function of RNAP is the transfer of the nucleotidyl motif firom the rNTP substrates to the hydroxyl at the 3 -end of the nascent RNA transcript. The nucleotidyl transfer reaction can be simplified as... 

Boffa, L.C., Walker, J., Chen, T.A., Sterner, R., Mariani, M.R., and Allfrey, V.G. (1990) Factors effecting nucleosome structure in transcriptionally active chromatin. Histone acetylation, nascent RNA and inhibitors of RNA synthesis. Eur. J. Biochem. 194, 811-823. 

DNA, since proximity eflfects demand that the DNA or nascent RNA closest to the histones at the point of disruption will be the polyanion for which those histones will preferentially reassociate. Ten Heggeler-Bordier et al. [95] have verified these observations. They used immuno-electron microscopy to determine what happens to histones after transcription with T7 RNA polymerase of a multi-nucleosomal template and also observed transfer to the nascent RNA. In contrast, Kirov et al. [96] have reported that no histones displace during transcription with this polymerase. However, as described above, transcriptional efficiency and ultimately histone displacement is not efficient in very low ionic strength conditions. 

Nascent RNA. The initial transcripts of RNA, before any modification or processing. 

Fig. 3. A transcription bubble. The DNA double helix Is unwound and RNA polymerase then synthesizes an RNA copy of the DNA template strand. The nascent RNA transiently forms an RNA-DNA hybrid helix but then peels away from the DNA which Is subsequently rewound into a helix once more.

The bacterial RNA polymerase has a subunit composition of ap>3 a, the a subunit being involved in correct initiation of transcription. Appropriate regulatory protein binding to the promoter region permits correct RNA polymerase binding, double-stranded DNA (dsDNA) unwinding and correct initiation of transcription. The dsDNA unwinds and the nascent RNA forms a transient RNA-DNA hybrid in the transcription bubble of unwound DNA that moves down the DNA. 

Mironov AS, Gusarov I, Raflkov R, Lopez LE, Shatalin K, Kreneva RA, Perumov DA, Nudler E. Sensing small molecules by nascent RNA a mechanism to control transcription in bacteria. Cell 2002 111 747-756. 

It is noteworthy that RNA polymerase lacks nuclease activity. Thus, in contrast with DNA polymerase, RNA polymerase does not correct the nascent polynucleotide chain. Consequently, the fidelity of transcription is much lower than that of replication. The error rate of RNA synthesis is of the order of one mistake per 10 or lO nucleotides, about 10 times as high as that of DNA synthesis. The much lower fidelity of RNA synthesis can be tolerated because mistakes are not transmitted to progeny. For most genes, many RNA transcripts are synthesized a few defective transcripts are unlikely to be harmful. 

How does this combination hairpin-oligo(U) structure terminate transcription First, it seems likely that RNA polymerase pauses immediately after it has synthesized a stretch of RNA that folds into a hairpin. Furthermore, the RNA-DNA hybrid helix produced after the hairpin is unstable because its rU-dA base pairs are the weakest of the four kinds. Hence, the pause in transcription caused by the hairpin permits the weakly bound nascent RNA to dissociate from the DNA template and then from the enzyme. The solitary DNA template strand rejoins its partner to re-form the DNA duplex, and the transcription bubble closes. 

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