Trans-acting siRNA


Posted by mmcmanus
presentation: 
evolution and conservation: 

Ta-siRNA activity was originally discovered in Arabidopsis (Peragine, A. et al) (Vazquez, F. et al). Since then, various groups have demonstrated the existence of various ta-siRNAs in both rice (Heisel, SE. et al) and maize (Williams, L. et al). While Arabidopsis contains several ta-siRNAs not found in other plants, tasiR-ARF is highly conserved in all of these systems. This indicates that, like miRNAs, ta-siRNAs (and tasiR-ARF in particular) have been used to regulate gene expression in plants since before the separation between the monocot and dicot lineages (Williams, L. et al). In 2006, David Bartel's group at MIT showed that, similar to Arabidopsis, ta-siRNAs derived from miR390 guided cleavage can be found in moss as well (see also Talmor-Neiman, M. et al). They argue that the use of dual miRNA recognition sites to guide ta-siRNA biogenesis is a mechanism that has been conserved for at least the last 400 million years (Axtell, MJ. et al).

significant partner RNAs or proteins: 

The biogenesis of ta-siRNAs requires RNA-dependent Polymerase 6 (RDR6), Suppressor of Gene Silencing 3 (SGS3), Dicer-like Protein 4 (DCL4) (Peragine, A et al) (Vazquez, F. et al) and possibly other Dicer-like proteins (Gasciolli, V. et al). A small subset of miRNAs guide the cleavage of TAS transcripts into ta-siRNAs:

miRNA directed cleavage of TAS gene transcripts requires a subset of the Argonaute proteins (Montgomery, TA. et al). The activity of ta-siRNAs also requires members of the RNA-induced Silencing Complex (RISC) (Peragine, A. et al) (Vazquez, F. et al).

mechanism of action: 

Once the 21-nt ta-siRNAs have been generated from a TAS gene, they act like other siRNAs and are incorporated into a RNA-induced Silencing Complex (RISC) where they guide the complex to cleave target mRNAs and thus repress translation (Peragine, A. et al) (Vazquez, F. et al). RISC complexes that incorporate ta-siRNAs must contain the appropriate Argonaute protein (of which there are 10 in Arabidopsis) to function properly. For instance, TAS3 derived tasiR-ARF requires Argonaute Protein 7 (AGO7) to cleave its target (Adenot, X. et al). Traditional siRNAs tend to target mRNAs with high sequence complementarity that originate from the same genomic location as the siRNA. Unlike these "traditional" siRNAs, but similar to miRNAs, ta-siRNAs bind to (and thus target the cleavage of) mRNAs with only partially identical sequences (Peragine, A. et al) (Vazquez, F. et al). Despite this, each ta-siRNA has a high target specificity (Allen, E. et al) This activity requires components of the miRNA pathway, but not components needed for heterochromatic RNAs, and thus links the once disparate siRNA and miRNA pathways (Vazquez, F. et al).

cellular functions: 

At least three different types of siRNAs are found in plants: ta-siRNA, nat-RNA and heterochromatic siRNA (Mallory, AC. and Vaucheret, H.). Unlike other siRNAs in plants, ta-siRNAs silence gene expression by acting in trans to cleave mRNAs with sequences partially complementary to their own (Peragine, A. et al). As such, the cellular function of each ta-siRNA is completely defined by its target and pattern of expression. So far, the ta-siRNAs generated from each TAS gene seem to have a common set of targets. The existence of four different families of TAS genes has been well established in Arabidopsis:

  • TAS1 derived ta-siRNAs regulate at least 5 different targets with currently unknown functions (Allen, E. et al).
  • TAS2 derived ta-siRNAs target 2 different Pentatricopeptide Repeat (PPR) proteins of disputed function (Allen, E. et al). There are around 450 PPR proteins in Arabidopsis, some of which may pay a role in RNA editing (Mach, J.).
  • TAS3 derived ta-siRNAs target three Auxin Response Factor (ARF) genes (ARF2, ARF3/ETT and ARF4) (Allen, E. et al)(Williams, L. et al). Auxins are signaling molecules that promote the vegetative development of Arabidopsis from a juvenile to adult stage. TAS3 ta-siRNAs (and tasiR-ARF in particular) inhibit this transition (Peragine, A. et al). Since the levels of tasiR-ARF, ARF3/ETT and ARF4 don't change during development, Hunter, C et al have suggested that these ta-siRNAs set the threshold at which leaves respond to Auxins. Recently, Chitwood, DH et al reported that tasiR-ARF can move intercellularly from the upper to the lower side of leaves to create what is essentially a morphogen gradient that patterns ARF3/ETT (see also Nogueira, FT. et al).
  • TAS4 derived ta-siRNAs target MYB transcripts (Rajagopalan, R. et al), though little work has been done on this locus. MYB genes are thought to control  functions as diverse as cell cycle, secondary metabolism and cell fate (Stracke, R. et al).
  • A TAS gene was also identified in moss (also named TAS4) and is thought to produce ta-siRNAs which target a putative transcription factor. While the activity of this target gene has not been characterized, its similarity to the AP2/EREBP transcription factor found in Arabidopsis suggests it may play a similar role in development (Talmor-Neiman, M. et al).

Computational analysis of the Arabidopsis genome indicates the potential existence of many other TAS genes (Chen, HM. et al), leaving open the possibility that a variety of new functions for ta-siRNAs may be uncovered in the future.

reviews: 
  1. Mallory, AC. and Vaucheret, H. Functions of microRNAs and related small RNAs in plants. Nature Genetics 38, S31 - S36 (2006). | PubMed
  2. Nogueira, FT. et al. Organ polarity in plants is specified through the opposing activity of two distinct small regulatory RNAs. Cold Spring Harb Symp Quant Biol. 71, 157-64 (2006). | PubMed
  3. Vaucheret, H. MicroRNA-dependent trans-acting siRNA production. Sci STKE. 2005(300), pe43 (2005). | PubMed
  4. Warkocki, Z. and Figlerowicz, M. [Trans-acting short interfering RNAs]. Postepy Biochem. 52: 253-9 (2006). | PubMed
  5. Willmann, MR. and Poethig, RS. Time to Grow up: the Temporal Role of smallRNAs in Plants. Curr Opin Plant Biol. 8, 548–552 (2005). | PubMed
  6. Xie, Z. and Qi, X. Diverse small RNA-directed silencing pathways in plants. Biochim Biophys Acta. 1779, 720-4 (2008). | PubMed
biogenesis: 

Each TAS gene so far identified produces a non-protein coding transcript that contains multiple ta-siRNAs within it. Each of these ta-siRNAs are lined up one after the other in both sense and anti-sense orientations. After transcription of the TAS gene, specific miRNAs pair with certain members of the Argonaute (AGO) protein family and bind to the single stranded RNA (ssRNA) at miRNA recognition sites (Montgomery, TA. et al). This specifies site specific cleavage of the primary TAS gene transcript at the beginning of the first ta-siRNA in the series and sets the phase for future processing (Yoshikawa, M. et al) (Allen, E. et al). A second, downstream miRNA-guided cleavage of the transcript is usually required for ta-siRNA biogenesis to continue as well (Axtell, MJ. et al). It has been proposed that ta-siRNA directed cleavage and phasing can also occur, though this is less well established and may be much more rare (Chen, HM. et al). At this point, Suppressor of Gene Silencing 3 (SGS3) protects the ssRNA from degradation (Elamayan, T. et al). Next, RNA-dependent Polymerase 6 (RDR6) produces a complementary strand, turning the transcript into double stranded RNA (dsRNA) (Yoshikawa, M. et al). Dicer-like Protein 4 (DCL4) then cleaves the dsRNA in 21-nt increments to generate mature ta-siRNAs (Yoshikawa, M. et al) (Xie, Z. et al) This is a partially redundant system in Arabidopsis, as the loss of one Dicer-like proteins can sometimes be compensated for by one of the three others (Gasciolli, V. et al).

references: 
  1. Adenot, X. et al. DRB4-dependent TAS3 trans-acting siRNAs control leaf morphology through AGO7. Curr Biol. 16, 927-32 (2006). | PubMed
  2. Allen, E. et al. microRNA-Directed Phasing during Trans-Acting siRNA Biogenesis in Plants. Cell 121, 207–221 (2005). | PubMed
  3. Axtell, MJ. et al. A Two-Hit Trigger for siRNA Biogenesis in Plants. Cell 127, 565-77 (2006). | PubMed
  4. Chen, HM. et al. Bioinformatic prediction and experimental validation of a microRNA-directed tandem trans-acting siRNA cascade in Arabidopsis. Proc Natl Acad Sci U S A. 104, 3318-23 (2007). | PubMed
  5. Chitwood, DH. et al. Pattern formation via small RNA mobility. Genes Dev. 23, 549-54 (2009). | PubMed
  6. de la Luz Gutiérrez-Nava, M. et al. Artificial trans-acting siRNAs confer consistent and effective gene silencing. Plant Physiol. 147, 543-51 (2008). | PubMed
  7. Elamayan, T. et al. A neomorphic sgs3 allele stabilizing miRNA cleavage products reveals that SGS3 acts as a homodimer. FEBS J. 276, 835-44 (2009). | PubMed
  8. Gasciolli, V. et al. Partially redundant functions of Arabidopsis DICER-like enzymes and a role for DCL4 in producing trans-acting siRNAs. Curr. Biol. 15, 1494–1500 (2005). | PubMed
  9. Heisel, SE. et al. Characterization of Unique Small RNA Populations from Rice Grain. PLoS ONE 3, e2871 (2008). | PubMed
  10. Hunter, C. et al. Trans-acting siRNA-mediated repression of ETTIN and ARF4 regulates heteroblasty in Arabidopsis. Development. 133, 2973-81 (2006). | PubMed
  11. Mach, J. Chloroplast RNA Editing by Pentatricopeptide Repeat Proteins. The Plant Cell 21, 17 (2009). | PubMed
  12. Matzke, M. and Scheid, OM. Epigenetic Regulation in Plants. In: Allis, CD., Jenuwein, T. & Reinberg, D. ed. Epigenetics. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press; 2007: 167-191. | Google Books | Amazon
  13. Montgomery, TA. et al. Specificity of ARGONAUTE7-miR390 interaction and dual functionality in TAS3 trans-acting siRNA formation. Cell 133, 128-41 (2008). | PubMed
  14. Peragine, A. et al. SGS3 and SGS2/SDE1/RDR6 are required for juvenile development and the production of trans-acting siRNAs in Arabidopsis. Genes Dev. 18, 2368–2379 (2004). | PubMed
  15. Rajagopalan, R. et al. A diverse and evolutionarily fluid set of microRNAs in Arabidopsis thaliana. Genes Dev. 20, 3407-25 (2006). | PubMed
  16. Stracke, R. et al. The R2R3-MYB gene family in Arabidopsis thaliana. Curr Opin Plant Biol. 4, 447-56 (2001). | PubMed
  17. Talmor-Neiman, M. et al. Identification of trans-acting siRNAs in moss and an RNA-dependent RNA polymerase required for their biogenesis. Plant J. 48, 511-21 (2006). | PubMed
  18. Vazquez, F. et al. Endogenous trans-Acting siRNAs Regulate the Accumulation of Arabidopsis mRNAs. Mol. Cell 16, 69-79 (2004). | PubMed
  19. Williams, L. et al. A database analysis method identifies an endogenous trans-acting short-interfering RNA that targets the Arabidopsis ARF2, ARF3, and ARF4 genes. Proc Natl Acad Sci U S A. 102, 9703-8 (2005). | PubMed
  20. Xie, Z. et al. DICER-LIKE 4 functions in trans-acting small interfering RNA biogenesis and vegetative phase change in Arabidopsis thaliana. Proc. Natl. Acad. Sci. USA 102, 12984–12989 (2005). | PubMed
  21. Yoshikawa, M. et al. A pathway for the biogenesis of trans-acting siRNAs in Arabidopsis. Genes Dev. 19, 2164–2175 (2005). | PubMed
example members: 

Four gene families have been identified in Arabidopsis that each produce a number of ta-siRNAs: TAS1, TAS2, TAS3 and TAS4. (Allen, E. et al). Within the TAS1 family, TAS1a, TAS1b and TAS1c are very similar in sequence and all produce the siR255 ta-siRNA among others (Allen, E. et al). One of the most famous ta-siRNAs is the trans-acting short-interfering RNA-auxin response factor (tasiR-ARF). Auxins are a class of signaling molecules that play a central role in plant development. TasiR-ARF targets the mRNA of three Auxin Response Factor (ARF) genes (ARF2, ARF3/ETT and ARF4) for degradation. (Williams, L. et al). TasiR-ARF is derived from the TAS3 gene (Allen, E. et al).

nomenclature: 

Trans-acting siRNA is frequently abbreviated as "ta-siRNA" and sometimes as "tasiRNA." Ta-siRNAs are frequently referenced in groups according to the Trans-acting siRNA (TAS) genes/loci from which they originate (e.g. "TAS1 derived ta-siRNAs"). Individual ta-siRNA names follow the convention for standard siRNAs: "siR" followed by a number (e.g. siR255).

potential as a tool: 

A variety of methods exist which use the endogenous posttranscriptional gene silencing pathway to repress gene expression in a targeted manner. Recently, a DuPont Crop Genetics Research group led by Robert Williams demonstrated that it was possible to co-opt the endogenous TAS genes in Arabidopsis to silence the expression of new target genes. By replacing the ta-siRNAs in the TAS1c gene with sequences from the Fatty Acid Desaturation 2 (FAD2) gene, the Williams group was able to reduce FAD2 activity down to levels observed in a null allele (de la Luz Gutiérrez-Nava, M. et al). Whether this silencing mechanism will work in non-plant systems is currently unknown, but it is tempting to imagine utilizing transgenic TAS genes as a method of delivering multiple ta-siRNAs to a single cell.

discovery: 

Credit for the discovery of ta-siRNAs is generally assigned to two groups who published near simultaneous papers in October of 2004. R. Scott Poethig's group at the University of Pennsylvania published a paper in Genes and Development describing how an Arabidopsis gene was "silenced posttranscriptionally in trans" by a family of siRNAs (Peragine, A. et al). Seven days later, Patrice Crété's group at the Université des Sciences et Technologies de Lillein in France published a paper in Molecular Cell that described a new set of siRNAs in Arabidopsis and layed out the basics of how they differed from other regulatory small RNAs. Crété's group dubbed this new class of siRNA "trans-acting siRNA" because "they direct hetero-silencing, repressing the expression of genes that bear little resemblance to the genes from which they derive" (Vazquez, F. et al). It is unclear whether Poethig and Crété were aware of each other's work prior to publication. In 2005, two groups demonstrated the important role miRNA guided cleavage played in generating ta-siRNAs and thus linked miRNA regulation to siRNA regulation (Yoshikawa, M. et al) (Allen, E. et al). This was expanded upon in 2006 when a paper published in Cell showed that dual miRNA cleavage sites were usually required for ta-siRNA biogenesis and constituted an evolutionarily conserved regulatory mechanism (Axtell, MJ. et al).

role in human disease: 

Whether ta-siRNAs exist in animals is currently unknown (Matzke, M. and Scheid, OM.). Unsuprisingly, ta-siRNAs have not been linked to any human diseases so far.