Abort! Abort!

Sometimes things go so wrong that it is just easier to start all over again. Bacteria have these situations too - it's not just us, humans! - and the central dogma of molecular biology (DNA replication, transcription and translation) is no exception.

In essence all the three steps of the central dogma share the very same basic topology: there is a message that gets read, there is a tool that reads it and there is a product. It looks like so:

Say, in the case of translation mRNA (the message) gets read by the ribosome (the tool) and protein (the product) is produced. And when things go wrong, there are three things you can abort: the message, the product and the tool. Let us see how it goes.

Replication

DNA polymerase (the tool) reads the DNA (the message) and produces DNA (the product). And when wrong nucleotide is incorporated, DNA polymerase can excise it and continue making the product using so called  proof-reading mechanism. Complete abortion of the growing DNA strand does not happen, and if mistake is done, it is done and you live with it. Surely, there are ways to fix it later (recombination and so on), but not on the spot, during the replication.

Transcription


RNA polymerases can proof-read too. However, many more things can be done. Special set of transcription factors, called GreA and GreB in bacteria and TFSII in eucaryotes, can activate intrinsic hydrolytic activity of the RNA polymerase and cleave off the growing product. Stalled complex is resolved and now we can try again.

Translation


First, there is a proof-reading mechanism, but rather than cutting off the mis-incorporated letter, GTP is hydrolyzed by GTPase EF-Tu which brings the aminoacyl-tRNA.

Second, if the mistake is done, and wrong amino acid was incorporated after all, bacterial class-1 release factors RF1 and RF2 become prone to peptide-release independent of the stop codon, thus removing the product (the growing protein chain). In mitochondria translational system is bacterial-like, but much more insane, and several (as many as 4 in humans!) class-1 release factors are present, with some of them lacking the ability to recognize the stop codon at all (ICT1, for example), and these resolve stalled ribosomal complexes by cutting off the peptide as well as their bacterial counterparts.

Third, bacterial toxins such RelE and the like are resolving ribosomal complexes by cutting the message (mRNA) rather than the product. Calling them toxins is rather misguiding, they are more of the rescue factors.

And lastly, eukaryotic translational factors Dom34 and Hbs1 (related to termination factors eRF1 and eRF3) are splitting the stalled ribosome into subunits, re-setting the tool.

So it seems the further we move from the DNA, the more dispensable the production complex becomes: in the case of DNA polymerases we have only proof-reading, RNA polymerases can do that and also cleave the message, and translational machinery can do it all: cutting the message (RelE), cutting the product (release factors) and resetting the tool by splitting the ribosome into subunits (Dom34 and Hbs).

References:

Borukhov S, Sagitov V, & Goldfarb A (1993). Transcript cleavage factors from E. coli. Cell, 72 (3), 459-66 PMID: 8431948

Toulmé F, Mosrin-Huaman C, Sparkowski J, Das A, Leng M, & Rahmouni AR (2000). GreA and GreB proteins revive backtracked RNA polymerase in vivo by promoting transcript trimming. The EMBO journal, 19 (24), 6853-9 PMID: 11118220

Pedersen K, Zavialov AV, Pavlov MY, Elf J, Gerdes K, & Ehrenberg M (2003). The bacterial toxin RelE displays codon-specific cleavage of mRNAs in the ribosomal A site. Cell, 112 (1), 131-40 PMID: 12526800

Orlova M, Newlands J, Das A, Goldfarb A, & Borukhov S (1995). Intrinsic transcript cleavage activity of RNA polymerase. Proceedings of the National Academy of Sciences of the United States of America, 92 (10), 4596-600 PMID: 7538676

Kassavetis GA, & Geiduschek EP (1993). RNA polymerase marching backward. Science (New York, N.Y.), 259 (5097), 944-5 PMID: 7679800

Richter R, Rorbach J, Pajak A, Smith PM, Wessels HJ, Huynen MA, Smeitink JA, Lightowlers RN, & Chrzanowska-Lightowlers ZM (2010). A functional peptidyl-tRNA hydrolase, ICT1, has been recruited into the human mitochondrial ribosome. The EMBO journal, 29 (6), 1116-25 PMID: 20186120

Shoemaker CJ, Eyler DE, & Green R (2010). Dom34:Hbs1 promotes subunit dissociation and peptidyl-tRNA drop-off to initiate no-go decay. Science (New York, N.Y.), 330 (6002), 369-72 PMID: 20947765

Atkinson GC, Baldauf SL, & Hauryliuk V (2008). Evolution of nonstop, no-go and nonsense-mediated mRNA decay and their termination factor-derived components. BMC evolutionary biology, 8 PMID: 18947425

Antonicka H, Ostergaard E, Sasarman F, Weraarpachai W, Wibrand F, Pedersen AM, Rodenburg RJ, van der Knaap MS, Smeitink JA, Chrzanowska-Lightowlers ZM, & Shoubridge EA (2010). Mutations in C12orf65 in patients with encephalomyopathy and a mitochondrial translation defect. American journal of human genetics, 87 (1), 115-22 PMID: 20598281

Zaher HS, & Green R (2009). Quality control by the ribosome following peptide bond formation. Nature, 457 (7226), 161-6 PMID: 19092806

mitochondrial mRNA UTRs: insanity, lunacy and absurd

mRNA in general have 5'UTR (untranslated region) followed by ORF (open reading frame) and then comes 3'UTR. Both UTRs regulate mRNAs stability, localization, translational efficiency. In eukaryotes cytozolic mRNAs have 3'UTR polyA, which regulates mRNA stability and translational efficiency, and 5'UTR which regulates efficiency of translation.

Mitochondrial mRNAs... it's a mess.

1) 3'UTRs are regulating localization (i.e. transport into the mitochondria, (Pattini 2003 PIMD 15376913)) and in mRNA stability (Gagliardi 2004 PIMD 15145579). Mechanisms of mRNA stabilization seem to be very, very different in plants (polyA destabilizes, like it does in bacteria!), yeast (no polyA at all, just like it is in yeast cytozolic mRNAs!) and mammals (polyA stabilizes, just like it does in eukaryotic cytozolic mRNAs!). 3 different approaches?! It is about as insane as it gets, really.

In mammals 3'UTRs are short (Ojala 1981 PIMD 7219536), unlike in plants and yeast, where 3' UTRs are long.

2) 5'UTRs are regulating translation (mRNA-specific enhancers of translation bind to them and thus regulate translation initiation) and localization within the mitochondria, thus coupling translation and insertion of the mRNA into the membrane. Most of the proteins that are translated in yeast mitochondria are intermembrane proteins involved in respiration and ATP production (review Towpik 2005 PIMD 16341268), thus coupling translation and insertion is a must. And indeed, when multisubunit complexes are assembled, translation of the individual subunits is geometrically coordinated (Naithani 2003 PIMD 12529447).

All this is well and good, but there are issues. Somehow investigation of 3' and 5'UTRs is a big thing in plant mitochondria, and not much is done nowadays with yeast or mammals. Or at least it is not easy to find. Second, no one tried systematically comparing UTRs from different organisms.

What I have dug out by now is this: in plants 5'UTRs are long, and there is a lot of experimental material here using 5'-RACE (Froner 2007 PIMD 17488843, Kuhn 2005 PIMD 15653634). In yeast - long 5'UTRs as well (review Costanzo 1990 PIMD 2088182), though much less studied experimentally. In mammals 5'UTRs are claimed to be short, at least in humans (Montoya 1981 PIMD 7219535). Here signals for mRNA-specific initiation enhancers are located within the ORF.

Is plant and yeast mitochondria translation radically different? Why did mammalian mitochondrial mRNA loose 5'UTR regulation? If yes, where is the watershed? What are the differences in the yeast+plants mitochondirial machinery vs mammalian?

Mammalian mitochondrial genome is super-streamlined, cutting corners where possible (review Attardi 1985 PIMD 3891661), so that could be a reason for the loss of 3' and 5'UTR. But what drives this minimization? Why trying THAT hard?

Experiments on mitochondrial translation seem to be done on different systems in different areas of research: yeast are used for identifying initiation enhancers and studying genetics and molecular biology of translation regulation, in plants 3' and 5' UTRs are extensively mapped, and in mammals using very, very simplified translational system (tRNA, IF2, IF3, EF-Tu and EF-G) some rudimentary biochemistry is done. This does scitsofrenic devision of labor is in keeping with the mitochondrial spirit indeed.

PS: mitochondrial ribosomes are also very, very strange.

Reviews:

Mitochondrial evolution: Karlberg 2003 PIMD 12728281
Mitochondrial and chloroplast translation: Gillham 1994 PIMD 7893142
Mitochondrial translation and desease: Perez-Martinez 2008 18991722
Plant mitochondrial translation: Binder 2003 PIMD 12594926, Hoffmann 2001 PIMD 11642360
Yeast mitochondrial translation: Costanzo 1990 PIMD 2088182, Dieckmann 1994 PIMD 8206703
Mammalian mitochondrial translation: Spremulli 2004 PIMD 15196894