Alternative polyadenylation in development

One of the things that is an open book is the true scope and physiological relevance of alternative polyadenylation.  A recent report in PNAS stirs this pot a bit (even if it leaves things still very much up in the air).  Briefly, this group has analyzed various large-scale gene expression repositories – ESTs, SAGE, and microarray – and found a tantalizing possible progression of 3′-UTR length during development.  Specifically, it seems as if global (or average) 3′-UTR length increases during the course of embryogenesis.  This change in the length of 3′-UTRs seems to be due to differential poly(A) site choice.  As I said, very tantalizing.

The abstract and brief commentary follows after the fold.

Where polyadenylation, siRNAs, and DNA methylation meet

It has become more apparent in recent years that the different aspects of gene expression – transcription initiation, transcription elongation, mRNA capping, splicing, and polyadenylation, transport of the mRNA to the cytoplasm, translation, and mRNA quality control – are rather extensively interconnected.  One corollary is that the polyadenylation complex, through various of its subunits, plays roles in various of these other processes.  This has been established for the most parts in mammalian and yeast models, but some recent work in plants is adding new and important variation to this theme.

A most recent of such studies has appeared online on PNAS.  This study, from the lab of Caroline Dean, reveals that the polyadenylation factor subunit FY (a homolog of the yeast protein Pfs2), acting in concert with the flowering regulator FCA, plays a crucial role in chromatin modifications that regulate the expression of the FLC gene.  Interestingly, this effect is not limited to just the FLC gene. Rather, other genes that are silenced by small RNA-mediated DNA methylation also require FY for this silencing.  This provocative finding seems to place FY in some sort of proximity to the small RNA-guided DNA methylation machinery, and may have some relevance to many aspects of transcription and mRNA quality control.

The abstract and citation follows. As always, enjoy.

Convergent transcription, polyadenylation, RNA editing, and regulation

One of the mechanisms by which polyadenylation may contribute to the regulation of gene expression (on paper, at least) involves gene pairs that are situated near each other and transcribed convergently.  In these instances, polyadenylation and transcription termination need to occur to prevent the production of RNAs that are anti-sense to the two members of the convergently-transcribed gene pair.  Overlapping transcripts could lead to the formation of double-stranded RNAs that could in turn trigger regulatory mechanisms, resulting in altered accumulation of the corresponding transcripts.

It is in this vein that a recent study from Gordon Carmichael’s lab at the University of Connecticut is of interest.  Briefly, these authors report that the early-to-late transition in gene expression in cells infected with the mouse polyoma virus is accomplished (at least in part) by a reduction in polyadenylation efficiency of the primary transcript encoding the so-called late genes.  Interestingly enough, this reduction in polyadenylation efficiency seems to be due to A-to-I editing of the region around the polyadenylation signal.  This editing in turn may be traced to an overlap of the early and late transcripts, such that double-stranded RNAs (the substrate for the A-to-I editing complex) that include the late polyadenylation signal are produced and edited before pre-mRNA processing occurs.

What we’re talking about

A recent paper from James Manley’s lab details a proteomic analysis of the polyadenylation complex.

In this study, the polyadenylation complex was purified using an affinity technique, attaching a functional polyadenylation signal to a series of MS2 coat protein binding sites (of course, all within the context of an RNA), incubating this RNA with Hela cell extracts, and purifying the RNA using a maltose binding protein-MS2 coat protein fusion. The protein components of the complex were characterized using the so-called Multidimensional Protein Identification Technology (MudPIT). In addition to the usual players* (CPSF, CstF, CFIm) were found proteins suggestive of links with transcription, splicing, and DNA repair. Curiously, the sole poly(A) polymerase found was not the canonical enzyme but an isoform first identified as an enzyme that adenylates the RNA present in the so-called Signal Recognition Particle (SRP). Moreover, readily detectable was a testis-specific CstF64 variant (CstF64-tau) as well as the canonical 64 kD subunit of CstF. CFIIm was largely absent.

Interesting stuff

While my yard is recovering from the ice, and I from today’s UK game, I thought I would toss out a few interesting abstracts that touch on important and contentious issues.  Peek beneath the fold and, as always, enjoy.

An interesting take on poly(A) signals

One interesting facet of RNA biology is the matter of the occurrence and function of structural RNA units within RNAs inside of living cells.  Excellent examples of these are the many so-called riboswitches, motifs that bind metabolites and alter the functionalities of RNAs in which they reside.  As more and more examples of RNAs with catalytic activities become known, questions naturally arise as to whether such activities might impact RNA function in vivo.  A recent study poses just such a question for what is among the simplest of known catalytic RNAs, namely the manganese-dependent ribozyme.  This enzyme consists of little more that a GAAA-UUU complex.  These two motifs, that need not be adjacent in the RNA, are expected to occur at a high frequency in natural RNAs.  A recent report indeed finds that RNAs that possess such motifs indeed may be Mn-dependent ribozymes.  The physiological signifiance of this finding is, in my opinion, a bit of an open issue.  But the possibilities are fascinating.  The last sentence of the abstract (that follows) is especially provocative.

Structure? Who needs structure?

One of the more interesting aspects of the polyadenylation complex is the nature of the protein-protein interactions that help shape the machinery. A recent study has revealed a fascinating side of this story. It involves the interaction between a subunit known as Fip1 (Fip = Factor Interacting with Poly(A) polymerase) and poly(A) polymerase (affectionately abbreviated as PAP). This study is remarkable for two reasons. For one, it dispels some previously-held notions about the functional significance of this interaction. In addition, it reveals that Fip1 is a member of a large class of proteins that share a common feature – namely, they do not have a rigidly-specified 3-dimensional structure in and of themselves.

If I could only remember ….

A recent essay described the process of cytoplasmic polyadenylation, and the functioning of the process in animal oocyte maturation and early development.  Cytoplasmic control of translation is important, not only in the oocyte, but in other settings as well.  One of these is in the neuron, where gene expression in repsonse to synaptic stimulation can be controlled by the activation of stored mRNAs in the cytoplasm.  While details of this control have remained poorly understood, a suggestive requirement (well, it’s suggestive when it comes to the general subject of this blog) of CPEB for memory in Drosophila lends itself to the hypothesis that the activation of stored mRNAs in the neuron involves cytoplamsic polyadenylation.

It is thus of interest to see that Drosophila homologue of Gld2 (the cytoplasmic poly(A) polymerase mentioned here) is required for long-term memory.  The abstract of the recent report follows.  As always, enjoy.

Cytoplasmic polyadenylation

The polyadenylation of mRNAs is usually thought of as a process that occurs in the nucleus, and indeed this is the cellular compartment in which pre-mRNA processing and polyadenylation does occur. However, mRNA polyadenylation is not restricted to the nucleus. Indeed, one of the more fascinating and important mechanisms that control gene expression during oogenesis and early development, stages in some organisms (such as animals) when the nucleus is not “active”, is mRNA polyadenylation. In these cases, the process occurs in the cytoplasm.

During oocyte development, a large population of maternally-encoded mRNAs are synthesized and stored for “use” in particular stages of development. These maternal mRNAs typically possess short poly(A) tails (20-40 nts) and are not available for translation (being “masked” by a complex of RNA-binding proteins). During oocyte maturation or following fertilization, these masked mRNAs become polyadenylated and thus activated for translation. This activation is a regulated process that helps to coordinate the ballet of gene expression attendant with meiotic maturation and early development. As such, it touches on many tangential phenomena (such as movement of stored mRNAs within the cell).

What is of particular interest for this blog is the nature of the mechanism that mediates polyadenylation in the cytoplasm. As indicated in the following figure, this mechanism includes some familiar players as well as some equally-intriguing partners.

Associations of alternative poly(A) sites with transposable elements

Previously, I discussed alternative polyadenylation, noting the extent (as many as 50% of human genes) of the phenomenon.  While there is some conservation of patterns of alternative poly(A) sites in animals, many of these events are species specific.  This suggests a relatively active evolutionary dynamic, one that shapes mRNAs in many ways.  A soon-to-be published study recently-published study in Nucleic Acids Research shed light on this dynamic, showing that species-specific poly(A) sites are often associated with transposable elements (TEs).  The abstract of the paper: