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Plants are remarkable for their amazing diversity of forms. Yet despite enormous differences in their appearance, all plants have a similar underlying structure. Each is composed of a central stem, secondary stems branching off from the main stem, and numerous leaves and/or flowers. All of these structures are derived from a single population of undifferentiated stem cells called the shoot apical meristem (SAM). The stem cells reside at the very tip of the SAM where they continuously divide and replenish themselves, enabling the plant to grow throughout its life. They also produce progeny cells that have the potential to become any type of organ: a leaf, a flower, or another meristem, and they control the overall size of the stem. Precise regulation of stem cell activity is essential for balancing tip growth with organ production, and the capacity to maintain a dynamic equilibrium of meristem cell populations is critical for plants to adapt to changing environments. Disruption of this equilibrium by excessive loss or accumulation of stem cells profoundly affects plant architecture and development (Carles and Fletcher 2003; Williams and Fletcher 2005). |
| Confocal image of ArabidopsisSAM |
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The goal of our research is to determine the molecular mechanisms that establish and maintain active stem cell reservoirs in shoot apical and floral meristems, using Arabidopsis thaliana as a model system. The approaches we take involve a combination of forward and reverse genetics, molecular biology and biochemistry. We are currently analyzing several stem cell maintenance pathways. Mutations in the Arabidopsis CLAVATA3 (CLV3) gene cause unregulated stem cell accumulation, yielding shoot apical meristems up to one thousand times larger than normal. CLV3 encodes a small polypeptide that is expressed only in the shoot and floral stem cells. We have demonstrated that CLV3 plays a key role in a negative feedback loop that controls stem cell fate in Arabidopsis shoot and floral meristems (Fletcher et al., 1999; Brand et al., 2000). Current work is directed towards identifying additional components of the CLV3 pathway, and factors that regulate CLV3 activity. |
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clv3-2mutant flower |
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In addition to the CLV3 pathway, we have identified the Arabidopsis ULTRAPETALA1 (ULT1) locus as an important negative regulator of shoot and floral stem cell activity. ult1 mutants have enlarged SAMs, and form flowers with many more sepals and petals than wild type flowers (Fletcher 2001). We cloned the ULT1 gene and found that it encodes a SAND domain putative transcription factor expressed throughout shoot and floral meristems (Carles et al. 2005). We have demonstrated that ULT1 acts as a critical temporal component of a pathway that terminates stem cell activity during the formation of the Arabidopsis flower. Along with two other factors, LFY and WUS, ULT1 activates the floral homeotic gene AGAMOUS (AG) at the proper time during floral development (Carles et al., 2004). AG in turn represses WUS in the center of the flower, terminating stem cell production. Experiments are underway to characterize the biochemical properties and downstream targets of ULT1 and a related protein, ULT2, and to identify additional components of the pathway. |
| ULT1mutant flower |
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Several lab projects have investigated the reciprocal interaction between the stem cell reservoir in the shoot meristem and the development of lateral organs such as leaves. Stem cell activity and lateral organ polarity are both controlled by a common small RNA regulatory pathway. The microRNA miR166 cleaves transcripts of three of its HD-ZIP transcription factor target genes in the stem cells, which in turn negatively regulate the level of WUS transcription of three of its HD-ZIP transcription factor target genes in the stem cells, which in turn negatively regulate the level of WUS transcription within the niche cells (Williams et al., 2005b). In a related study, the lab used a database analysis approach to identify a novel small RNA species called tasiR-ARF, one of the first representatives of a recently defined class of small RNAs called trans-acting siRNAs (Williams et al. 2005a). tasiR-ARF negatively regulates three auxin response genes that are required for shoot maturation and the specification of lateral organ polarity, and indirectly down-regulates miR166 activity.
We have also determined that the BTB/POZ domain putative transcriptional regulators BOP1 and BOP2 negatively regulate stem cell fate and promote lateral organ differentiation (Ha et al, 2004). The BOP pathway converges with the ARF and miR166 pathways to establish lateral organ polarity (Ha et al. 2007), with BOP promoting adaxial identity by up-regulating HD-ZIP gene expression on the top side of the organs and miR166 promoting abaxial identity by cleaving HD-ZIP gene transcripts on the bottom side. Together, these studies are building an integrated picture of how stem cell maintenance and lateral organ formation are coordinated throughout plant development.
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| Wild-type Arabidopsis flower |
