| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
|
Plant Physiology 138:565-566 (2005) © 2005 American Society of Plant Biologists Small RNAs and Arabidopsis. A Fast Forward LookCenter for Gene Research and Biotechnology, Oregon State University, Corvallis, Oregon 97331
First, let's take a quick look back to a few formative events leading to the blur of activity that now defines the small RNA field. In 1993, Victor Ambros and colleagues (Lee et al., 1993 I feel at risk, therefore, projecting a future outlook for this fast-moving field. Statements concerning the most important questions to address and the technology needed to answer these questions may be entirely obsolete by the time this article appears in print. Nevertheless, I offer the following four areas as fertile ground for discovery in plant small RNA biology.
A large proportion of plant miRNA families, and some trans-acting (ta) siRNAs, function as negative regulators of mRNAs coding for transcription factors with roles in development (Jones-Rhoades and Bartel, 2004  ). For example, miR160 and miR167 each target clades within the auxin response factor family of transcription factors. Several others miRNA target genes encode factors controlling meristem function and patterning. Disruption of negative regulation by miRNAs, through mutation of target sites or miRNA sequences, frequently results in developmental phenotypes (Baulcombe, 2004). However, the mechanisms whereby miRNAs and ta-siRNAs are integrated into developmental or signaling pathways are understood poorly. To what extent do these small RNAs trigger decisive or committed steps in cell differentiation? Do they also function as constitutive negative regulators to direct or reinforce the "off" state of target genes? How are miRNA genes controlled?
It is clear that dynamic, cell-specific imaging of expression or activity patterns of specific miRNAs and ta-siRNAs, target mRNAs, and downstream genes over time is needed to place these small RNAs into spatial and temporal contexts during development. Two major types of resources are either under development or in early deployment to address these problems. The first is sensor technology, in which fluorescent proteins serve to reveal cell-specific patterns of miRNA gene expression and targeting activity. For example, sensor constructs containing miRNA target sequences are particularly useful as reporters of miRNA activity in situ (Parizotto et al., 2004
An early eukaryote likely possessed genes encoding a repertoire of RNAi factors, including Dicer-like (DCL), Argonaute, and RNA-dependent RNA polymerase proteins. The DCL protein contained two RNaseIII-like domains for double-stranded RNA processing, whereas the Argonaute protein likely possessed an RNaseH-like domain. Families for each of these three factors expanded in plants after divergence from other eukaryotic lineages, providing genes for specialization of miRNA, ta-siRNA, heterochromatin-associated RNAi, and antiviral RNAi pathways (Baulcombe, 2004 ). However, functions still remain to be assigned to the majority of the 10 Argonaute members in Arabidopsis (Arabidopsis thaliana) and some of the DCL factors. The Argonautes are particularly interesting. Functions in miRNA and siRNA-directed target cleavage have been assigned for only two family members (AGO1 and AGO4), while a third (ZIPPY) was functionally assigned as a factor in juvenile-to-adult phase transitions (Baulcombe, 2004). Why are there so many more AGO factors than known miRNA- and siRNA-dependent biochemical pathways? My guess is that there is far more pathway specialization than we currently realize.
In addition to small RNA biogenesis and effector factor families, the small RNA-generating loci themselves belong to gene families and are still evolving. Sequence duplication events resulting in foldback or hairpin transcripts appear to be one mechanism whereby new small RNA regulators with novel specificity can arise (Allen et al., 2004
Some of the best work to define biochemical components and activities associated with small RNA formation and effector pathways comes from analysis of fly extracts and other animal-based in vitro systems (Bartel, 2004 ). Relatively little plant RNAi biochemistry has progressed to the point of publication. While there are clearly some interesting biochemical issues that need to be addressed in plants, such as how different DCLs catalyze formation of different classes of small RNAs, there are many technical barriers that limit progress. Analysis of complexes from wild-type and mutant Arabidopsis plants or cells through immunoprecipitation or other affinity tags may offer a path around some of these barriers.
Another interesting set of questions concerns how RNAi complexes integrate with other cellular processes. For example, while it is clear that a set of specialized RNAi components (DCL3, RDR2, AGO4) participate in heterochromatin formation at many loci (Zilberman et al., 2003
Among the least understood areas of RNAi biology in plants concerns the intercellular mobility of small RNAs and small RNA-containing complexes. We have known for nearly a decade that sequence-specific information during RNA silencing is transported over short (cell-to-cell) and long (phloem-dependent) distances (Baulcombe, 2004 ). The antiviral response appears to depend, in part, on systemic transport of RNAi signals. However, the mechanistic basis of this pathway as well as the relevance of transport of active miRNAs and siRNAs during growth and development remain ill defined. Genetic data supporting identification and function of components of these transport pathways are absent. Clearly, creative genetic screens for informative mutants with specific defects in the RNA transport system(s) are desperately needed. In addition to these fundamental areas, the plant-based biotechnology industry is poised to deliver novel products that involve small RNAs. The predictability and broad utility of directed silencing using available RNAi technology and the potential of other technologies that deploy different small RNA pathways suggest that RNAi-conditioned phenotypes may proliferate in agricultural products. The limitations for industry appear more related to social forces rather than scientific potential.
www.plantphysiol.org/cgi/doi/10.1104/pp.104.900156. * E-mail carrington{at}cgrb.oregonstate.edu; fax 541–737–3045.
Allen E, Xie Z, Gustafson AM, Sung GH, Spatafora JW, Carrington JC (2004) Evolution of microRNA genes by inverted duplication of target gene sequences in Arabidopsis thaliana. Nat Genet 36: 1282–1290[CrossRef][ISI][Medline] Bartel D (2004) MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 116: 281–297[CrossRef][ISI][Medline] Baulcombe D (2004) RNA silencing in plants. Nature 431: 356–363[CrossRef][Medline]
Birnbaum K, Shasha DE, Wang JY, Jung JW, Lambert GM, Galbraith DW, Benfey PN (2003) A gene expression map of the Arabidopsis root. Science 302: 1956–1960
Hamilton AJ, Baulcombe DC (1999) A species of small antisense RNA in posttranscriptional gene silencing in plants. Science 286: 950–952
Herr AJ, Jensen MB, Dalmay T, Baulcombe DC (2005) RNA polymerase IV directs silencing of endogenous DNA. Science 308: 118–120 Jones-Rhoades MW, Bartel DP (2004) Computational identification of plant microRNAs and their targets, including a stress-induced miRNA. Mol Cell 14: 787–799[CrossRef][ISI][Medline] Lee RC, Feinbaum RL, Ambros V (1993) The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell 75: 843–854[CrossRef][ISI][Medline] Onodera Y, Haag JR, Ream T, Nunes PC, Pontes O, Pikaard CS (2005) Plant nuclear RNA polymerase IV mediates siRNA and DNA methylation-dependent heterochromatin formation. Cell 120: 613–622[CrossRef][ISI][Medline]
Parizotto EA, Dunoyer P, Rahm N, Himber C, Voinnet O (2004) In vivo investigation of the transcription, processing, endonucleolytic activity, and functional relevance of the spatial distribution of a plant miRNA. Genes Dev 18: 2237–2242 Xie Z, Johansen LK, Gustafson AM, Kasschau KD, Lellis AD, Zilberman D, Jacobsen SE, Carrington JC (2004) Genetic and functional diversification of small RNA pathways in plants. PLoS Biol 2: 642–652
Zilberman D, Cao X, Jacobsen SE (2003) ARGONAUTE4 control of locus-specific siRNA accumulation and DNA and histone methylation. Science 299: 716–719 This article has been cited by other articles:
|
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| ASPB Publications | PLANT PHYSIOLOGY | THE PLANT CELL | |
|---|---|---|---|
TOP
INTEGRATION OF SMALL RNA...
EVOLUTION AND SPECIALIZATION OF...
