Expression of a Micro-protein

Yu, Xiang; Warner, Jonathan R.

doi:10.1074/jbc.m103772200

Cited by 41 publications

(34 citation statements)

References 33 publications

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“…The RPL41A mRNA from Saccharomyces cerevisiae (encoded by YDL184C) was used because the 59 end is mapped and the mRNA is abundant (50 copies per cell) (Yu and Warner 2001). RPL41A coding sequence is closely related to RPL41B, yet these mRNAs differ substantially in their UTR sequences, making it possible to develop a detection strategy specific for RPL41A (Yu and Warner 2001).…”

Section: Resultsmentioning

confidence: 99%

A quantitative assay for measuring mRNA decapping by splinted ligation reverse transcription polymerase chain reaction: qSL-RT-PCR

Blewett¹,

Coller²,

Goldstrohm³

2011

RNA

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The degradation of messenger RNA is a critical node of gene regulation. A major pathway of mRNA decay is initiated by shortening of the poly(A) tail, followed by removal of the 59 cap structure (decapping) and subsequent degradation. Decapping is an important determinate in the destruction of many transcripts. Detailed kinetic analysis of in vivo decapping rates is necessary to understand how this step is regulated. Importantly, the product of decapping is recalcitrant for investigation, in part due to its transient nature. As such, little in vivo kinetic information is available. Here we report the development of an assay that measures decapping of mRNAs by combining splinted ligation and quantitative RT-PCR (qSL-RT-PCR). We apply this method to determine the decapping rate constant for a natural mRNA in vivo for the first time. The qSL-RT-PCR assay may be adapted for use on any mRNA, providing a new tool to study regulation of mRNA decay.

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Section: Resultsmentioning

confidence: 99%

A quantitative assay for measuring mRNA decapping by splinted ligation reverse transcription polymerase chain reaction: qSL-RT-PCR

Blewett¹,

Coller²,

Goldstrohm³

2011

RNA

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“…In this context, it is interesting to note that the smallest naturally occurring eukaryotic ORF that produces significant levels of protein product comprises 24 codons (Yu and Warner 2001). We have described a set of human b-globin gene nonsense mutations at codons 5, 15, and 17 that are not committed to NMD (Romão et al 2000).…”

Section: Discussionmentioning

confidence: 99%

The canonical UPF1-dependent nonsense-mediated mRNA decay is inhibited in transcripts carrying a short open reading frame independent of sequence context

Silva¹,

Pereira²,

Morgado³

et al. 2006

RNA

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Nonsense-mediated mRNA decay (NMD) is a surveillance mechanism that degrades mRNAs carrying premature translation termination codons. Generally, NMD is elicited if translation terminates >50-54 nucleotides (nt) upstream of an exon-exon junction. We have previously reported that human b-globin mRNAs carrying 59-proximal nonsense mutations (e.g., b15) accumulate to normal levels, suggesting an exception to the ''50-54-nt boundary rule.'' In the present report, we demonstrate that the strength of the UPF1-dependent NMD of mutant b-globin mRNAs is specifically determined by the proximity of the nonsense codon to the initiation AUG. This conclusion is supported by a parallel effect of the short ORF size on NMD of nonsense-containing a-globin mRNAs. To determine whether the short-ORF effect on NMD response is conserved in heterologous transcripts, we assessed its effects on a set of b-globin/triosephosphate isomerase (TPI) hybrid mRNAs and on the TPI mRNA. Our data support the conclusion that nonsense mutations resulting in a short ORF are able to circumvent the full activity of the canonical UPF1-dependent NMD pathway.

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“…L41 is a unique protein, with only 25 amino acids, of which 17 are arginine or lysine, highly conserved throughout the eukaryotes, and occupying an intimate location at the interface between the 60S and 40S subunits (Jenner et al 2012). With such a short ORF, the mRNA encoding L41 is translated by only one ribosome at a time (Yu and Warner 2001). Furthermore, stretches of basic amino acids are translated inefficiently, perhaps because of their difficulty in traversing the ribosome "tunnel" (Charneski and Hurst 2013).…”

Section: Stoichiometry Of Mrnas Encoding Rpsmentioning

confidence: 99%

Ribosome-omics of the human ribosome

Gupta

Warner

2014

RNA

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The torrent of RNA-seq data becoming available not only furnishes an overview of the entire transcriptome but also provides tools to focus on specific areas of interest. Our focus on the synthesis of ribosomes asked whether the abundance of mRNAs encoding ribosomal proteins (RPs) matched the equimolar need for the RPs in the assembly of ribosomes. We were at first surprised to find, in the mapping data of ENCODE and other sources, that there were nearly 100-fold differences in the level of the mRNAs encoding the different RPs. However, after correcting for the mapping ambiguities introduced by the presence of more than 2000 pseudogenes derived from RP mRNAs, we show that for 80%-90% of the RP genes, the molar ratio of mRNAs varies less than threefold, with little tissue specificity. Nevertheless, since the RPs are needed in equimolar amounts, there must be sluggish or regulated translation of the more abundant RP mRNAs and/or substantial turnover of unused RPs. In addition, seven of the RPs have subsidiary genes, three of which are pseudogenes that have been "rescued" by the introduction of promoters and/or upstream introns. Several of these are transcribed in a tissue-specific manner, e.g., RPL10L in testis and RPL3L in muscle, leading to potential variation in ribosome structure from one tissue to another. Of the 376 introns in the RP genes, a single one is alternatively spliced in a tissue-specific manner.

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Expression of a Micro-protein

Cited by 41 publications

References 33 publications

A quantitative assay for measuring mRNA decapping by splinted ligation reverse transcription polymerase chain reaction: qSL-RT-PCR

A quantitative assay for measuring mRNA decapping by splinted ligation reverse transcription polymerase chain reaction: qSL-RT-PCR

The canonical UPF1-dependent nonsense-mediated mRNA decay is inhibited in transcripts carrying a short open reading frame independent of sequence context

Ribosome-omics of the human ribosome

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