Putting U in the tail of mRNA

mRNA, the gene transcript that awaits translation, is made of many nucleotides of RNA. These RNA nucleotides can be modified and these modifications can have functional consequences that alters the lifespan of mRNA, its localisation or translation efficiency. Modifications can fall into one of two categories; either nucleotides can be individually targeted or several nucleotides can be added to the 3’end, often referred to as tailing.  mRNA typically contains a poly(A) tail at its 3’end, a string of adenine (A) residues, which aids export of mRNA from the nucleus and its translation in the cytoplasm of a cell. The length of the poly(A) tail varies between different mRNAs and during the life of a single mRNA. In addition to the variation in poly(A) length, it is becoming ever more apparent that uridylation, tailing of mRNA with U residues, plays an important role. Knowing that U residues can be added to the end of mRNA, this raises two key questions; what adds the U residues and what is its function?

TUTases

So, what adds the uridine residues to the terminal of the mRNA… the terminal uridylyltransferases (TUTases) of course! The TUTases are a sub-group of the pol β superfamily. Interestingly, the TUTases all contain long intrinsically disordered regions (IDRs) that are known to mediate protein-RNA interactions. Since IDRs are also flexible this may enable TUTases to recognise a diverse range of RNA substrates. Moreover (as discussed in a previous blog https://asheekeyscienceblog.com/2017/09/09/get-transcribed-in-style-the-ctd-that-everyones-talking-about/ ), IDRs are also enriched in phase separated droplets such as P-bodies and stress granules where mRNAs are thought to be stored, which suggests TUTases may localise the mRNA in the cell too1.

The U in decay?

The big clue for the role of uridylation in mRNA processing came when it was observed that uridylated mRNAs accumulated when mRNA degradation components were mutated. Moreover, the half-life of the urg1 mRNA increased when a TUTase (Cid1) was deleted. Both these results strongly support a role of uridylation in promoting mRNA degradation2.

Before I go into the nitty gritty of what is thought to occur on mRNA uridylation, it is important to be familiar with the components involved in mRNA degradation. The first step of degradation is the recruitment of deadenylases that remove A residues from the poly(A) tail – CCR4-NOT and Pan2/Pan3 are two complexes with deadenylation activity. Reducing the poly(A) tail reduces the stability of mRNA since it prevents the poly(A) binding protein (PABP) from binding. In addition, deadenylation recruits decapping proteins (Dcp1/Dcp2) that remove the 5’cap. Cap removal allows access of 5’ exonucleases (proteins that degrade mRNA from the 5’end) such as XRN1 destroying the mRNA. The basic outline of this degradation pathway can be seen in Figure 1.

 

degradation
Figure 1: mRNA decay

 

So how does uridylation fit into this pathway…?

A good place to start is with histone mRNAs. Histones are proteins that get wrapped around by DNA to compact DNA in the nucleus, but don’t get bogged down by its function… Histone mRNAs differ from most since they do not have a poly(A) tail. Uridylation of the 3’ end of the mRNA is a key trigger for degradation. The uridine residues act as a mini landing-pad for the Lsm1-7 complex. Lsm1-7 has decapping activity promoting 5’cap removal and mRNA degradation by the 5’exonucleases. The Lsm complex can also recruit the decapping factors which is what is thought to occur for other mRNAs (Figure 2).

 

decay u
Figure 2: Uridylation in mRNA degradation

 

Uridylation and deadenylation thus appear to play redundant roles in the recruitment of decapping enzymes. Can they coordinate together? More importantly, if they do, is a link between poly(A) tail length and uridylation?

Much research has already considered this question. In some species of yeast, small flowering plants and in humans it appears that deadenylation precedes uridylation. In humans, for example, the TUTases, TUT4/TUT7 have a preference for binding short poly(A) tails (~20 A’s) – at lengths greater than 25 A’s the tail is protected by the PABP. However, this is by no means the case for all. How widespread and whether either of these pathways dominate in a species/tissues/gene-specific manner is not yet certain. Nevertheless, it supports the view that mRNA turnover is a crucial cellular process that needs to be effectively performed (1).

Mammalian oocytes

Experiments to understand the importance of mRNA tail modifications have tended to opt for oocytes. In the early stages of development there is a lack of transcription and so instead to regulate gene expression, mRNA stability and translation regulation become more dominant. mRNA transcripts come from the maternal oocyte which are thus a good experimental model to test the complexities in tails.

A recent study published in Nature have looked at the role of uridylation in regulating this maternal ‘transcriptome’ in early mouse development 3. They discovered that TUT4 and TUT7 through the addition of U’s to different mRNA transcripts signal the mRNA for degradation effectively acting as ‘sculptors of the transcriptome’. Interestingly, deletion of TUT4 &7 in oocytes had huge effect causing accumulation of mRNAs with short poly(A) tails whilst depletion of these TUTases in somatic (“normal”) cells had a minor effect on gene expression. This greater importance for tail regulation in oocytes in early development compared to somatic cells, correlates well with the current understanding that poly(A) tail length is also more important for translational regulation in early development. It would be interesting to characterise these events on specific transcripts to further our knowledge into the tight regulation of translation in early development!

Where have we seen U before?

mRNAs are not the only nucleic acids to be uridylated; U residues are also known to be added to several other classes of non-coding RNA including miRNA and snRNA. U residues are also used in trypanosomes for mRNA editing; a diverse yet fascinating additional role for U’s.

Hopefully this has got U up to grips with uridylation.

Further reading

  1. Scheer, H., Zuber, H., De Almeida, C. & Gagliardi, D. Uridylation Earmarks mRNAs for Degradation… and More. Trends in Genetics 32, 607–619 (2016).
  2. Rissland, O. S. & Norbury, C. J. Decapping is preceded by 3′ uridylation in a novel pathway of bulk mRNA turnover. Nat. Struct. Mol. Biol. 16, 616–623 (2009).
  3. Morgan, M. et al. MRNA 3′ uridylation and poly(A) tail length sculpt the mammalian maternal transcriptome. Nature 548, 347–351 (2017).

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