CEPCEB Members
Xuemei Chen
Professor
Botany & Plant Sciences
University of California
Riverside, CA 92521
Phone: (951) 827-3988
Fax: (951) 827-4294
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| | Background
I received my B.S. degree from Beijing University in China in 1988 and joined the Ph.D. program in biochemistry at Cornell University in 1989. I did my Ph.D. research in the lab of Dr. David Stern at the Boyce Thompson Institute for Plant Research, where I studied posttranscriptional regulation of chloroplast gene expression in the unicellular alga Chlamydomonas. During this time, I developed a strong interest in patterning mechanisms that underlie the development of multicellular organisms. After receiving my Ph.D. in 1995, I joined Dr. Elliot Meyerowitz’s lab at California Institute of Technology to study molecular mechanisms that pattern the flower in Arabidopsis thaliana.
I became an Assistant Professor at the Waksman Institute at Rutgers University in 1999, where my lab continued to study mechanisms governing flower development and where we carried out pioneering work on plant microRNA biogenesis and function. During my tenure review at Rutgers University in 2005, I was chosen to receive the Board of Trustees Research Fellowship for Scholarly Excellence.
I moved to UC Riverside as an Associate Professor in the Department of Botany and Plant Sciences in 2005. Here, we continue to pursue our two research interests, floral patterning and small RNA biology.
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Cell Fate Specification in Floral Patterning
The Arabidopsis flower is a great model to dissect developmental mechanisms underlying patterning. The flower comes from a group of undifferentiated cells known as the floral meristem. A series of cell fate specification events occurs within the floral meristem during flower development. The stem cells in the center of the floral meristem divide to produce daughter cells, some of which remain stem cells while others are displaced to the periphery of the meristem to become floral organ primordia. The floral meristem puts out four types of organ primordia successively in rings, or whorls. The first and second whorl primordia become sepals and petals, respectively, which are also known as perianth organs. The third and fourth whorl primordia become stamens and carpels, respectively, which are reproductive organs. Upon production of the carpel primordia, the stem cells in the floral meristem are terminated such that no more floral organs are generated. Therefore, floral patterning involves the temporal regulation of floral stem cells and the specification of floral organ identity among other patterning events.
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| Figure 1. Arabidopsis flowers of various genotypes. (A) wild type. (B), hua1-1 hua2-1. (C) hua1-1 hua2-1 hen1-1. (D) hua1-1 hua2-1 hen3-1. (E) hua1-1 hua2-1 hen4-1. (F) hua1-1 hua2-1 paused 1 (hen5-1). The arrows indicate third whorl organs-stamens in wild type and hua1-1 hua2-1 and petals in the others. |
Molecular genetic analyses by many labs led to the discovery of four classes of genes (the so-called A, B, C, and E genes) encoding transcription factors that act in combination to specify the four floral organ identities. While the class E genes act in all four whorls, the A, B, and C genes each acts in two adjacent whorls: A in whorls 1+2, B in whorls 2+3, and C in whorls 3+4. The unique composition of the ABCE genes in each whorl specifies the identity of the whorl: A+E for sepal, A+B+E for petal, B+C+E for stamen, and C+E for carpel. Another important genetic interaction is the antagonism between the A and C genes, which helps restrict them to their normal domains of activity. In the absence of A function, C function expands into the outer two whorls to convert perianth organs into reproductive organs. In the absence of C function, A function expands into the inner two whorls to convert reproductive organs into perianth organs. In addition to its role in specifying reproductive organ identities, the class C gene AGAMOUS (AG) also plays a key role in the timely termination of floral stem cells. In an ag loss-of-function mutant, floral stem cells continue to put out floral organs to result in a flowers-within-flower phenotype.
We have been interested in dissecting the AG pathway that specifies reproductive organ identities and regulates the termination of floral stem cells. As a postdoctoral fellow in Dr. Elliot Meyerowitz’s lab, I performed a sensitized genetic screen in the weak ag-4 mutant background to isolate mutations that further compromise the AG pathway. Mutations in two new genes, HUA1 and HUA2, enhance the ag-4 defect, suggesting that HUA1 and HUA2 play a role in the AG pathway. We took advantage of the weak phenotype of the hua1 hua2 double mutant and performed another genetic screen to isolate mutations that enhance the hua1 hua2 phenotype such that the flowers resemble ag mutant flowers. Mutations in five new genes, which we named HUAENHANCER (HEN)1-5, were isolated (Figure 1). The ag-like phenotypes of the hua1hua2 hen mutants suggest that the HEN genes all play a role in the AG pathway in flower development. We cloned the five HEN genes as well as HUA1 by map-based cloning.
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| Figure 2. miR172 in flower development. ap2-2 (A) and 35S::M1R172 (B) have almost identical floral phenotypes. The arrows in A and B indicate carpels that replace sepals. ag-1 (C) and 35S::AP2m3 (D) have similar floral defects. E-F, in situ hybridization to determine miR172 localization (purple color) in young flowers. (E) miR172 is present in stages 1-2 floral meristems (FM) but absent from the inflorescence meristem (IM). (F) miR172 is enriched in the inner two floral whorls in a stage 7 flower. The numbers represent the positions of the floral whorls. Scale bars, 50 μm. |
Intriguingly, with the exception of HEN3 (Wang and Chen, 2004) [PDF], all HUA and HEN genes encode proteins with implicated cellular functions in RNA metabolism, suggesting that posttranscriptional mechanisms govern floral patterning. In particular, we found that HUA1, HUA2, HEN2, and HEN4 promote AG expression by preventing pre-mature transcription termination within the second intron of AG (Cheng et al., 2003) [PDF]. HEN1 encodes a novel protein required for the normal accumulation of microRNAs (Park et al., 2002) [PDF] and small interfering RNAs (siRNAs) (Boutet et al., 2003) [PDF]. Furthermore, we demonstrated that a microRNA, miR172, whose biogenesis requires HEN1, plays a key role as a repressor of the class A gene APETALA2 (AP2) (Chen, 2004) [PDF]. miR172 is specifically present in floral meristems (Figure 2 ). Mis-expression of MIR172 genes with the 35S promoter results in ap2 loss-of-function phenotypes (Figure 2 ). Mis-expression of AP2m3, a miR172-resistant version of AP2 cDNA but not wild-type AP2 cDNA, results in flowers that resemble ag loss-of-function mutants (Figure 2 ). Our current and future research involves the definition of the role of miR172 in flower development in relation to the AG-AP2 antagonistic pair and the further dissection of the mechanisms underlying the regulation of floral stem cells.
Small RNA Metabolism and Function
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| Figure 3. A schematic diagram of miRNA biogenesis in Arabidopsis. The precursor-miRNA (pre-miRNA) is processed by DCL1 to a duplex of the miRNA and its antisense strand miRNA*. HEN1 methylates the 3' terminal nucleotide in each strand of the duplex. The methylation occurs on the ribose of the terminal nucleotide. The methylated miRNA is incorporated into a protein complex names RISC, which contains an argonaute protein (AGO). The role of HYL1 in miRNA biogenesis is unknown. |
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miRNAs and siRNAs are 21-24 nt small RNAs that serve as sequence-specific regulators of protein coding genes. The small RNAs can act at multiple levels, such as chromatin modification, RNA cleavage, and translational repression.
Our interest in small RNA metabolism stemmed from the desire to understand the biochemical function of HEN1, a gene isolated from our genetic screen on flower development. We found that HEN1 plays a general role in the biogenesis of miRNAs and siRNAs (Park et al., 2002 [PDF]; Boutet et al., 2003 [PDF]). Most miRNAs and siRNAs are reduced in abundance and heterogeneous in size in hen1 mutants. The presence of a putative methyl transferase motif in the HEN1 protein prompted us to examine the potential function of HEN1 as a small RNA methyltransferase. We demonstrated that in vitro HEN1 methylates duplexes of small RNAs that mimic the products of Dicer-mediated cleavage of small RNA precursors (Figure 3) (Yu et al., 2005) [PDF]. HEN1 adds a methyl group to the 3’ terminal nucleotide of the small RNA. In addition, small RNAs do carry one methyl group on their 3’ terminal nucleotides in vivo (Yu et al., 2005). In hen1 mutants, the small RNAs lack the methyl group, suggesting that HEN1 is responsible for the methylation of small RNAs in vivo (Yu et al., 2005). Therefore, we have uncovered a new step in plant small RNA metabolism (Figure 3).
What is the function of small RNA methylation? In the absence of methylation, small RNAs become heterogeneous in size in vivo. We found that the small RNAs have 3’ extensions composed of mainly uridine residues (Li et al., 2005) [PDF]. This suggests that an as yet unknown enzyme targets the unmethylated small RNAs and that at least one function of the methylation is to protect the small RNAs from this undesirable activity.
Our current and future interests in small RNA metabolism center on the identification of the unknown uridylation activity and on understanding the role of small RNA methylation in the context of RNA silencing.
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Selected Publications (Bibliography page)
Current Laboratory Personnel
- Manu Argawal, postdoctoral fellow since Sept 2005.
- Julien Curaba, postdoctoral fellow since Feb 2005.
- YunJu Kim, graduate student since July 2005.
- Bin Yu, postdoctoral fellow since March 2004.
- Zhiyong Yang, postdoctoral fellow since Sept 2003.
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