This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Related articles in Plant Cell Similar articles in this journal Similar articles in ISI Web of Science Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Eckardt, N. A. Search for Related Content PubMed Articles by Eckardt, N. A. Agricola Articles by Eckardt, N. A.

IN THIS ISSUE

A Role for APETALA2 in Maintenance of the Stem Cell Niche

neckardt{at}aspb.org

Meristems maintain a small pool of undifferentiated stem cells that provides for the continual supply of new cells necessary for postembryonic organ formation and the growth of an organism. Meristems perform a finely tuned balancing act between maintenance of the stem cell niche and promotion of some daughter cells to differentiate and form organ primordia. In Arabidopsis, the stem cells in the shoot apical meristem (SAM) are located in three outermost cell layers of the central region of the apex. Stem cells are maintained in an undifferentiated state by the activity of a small group of cells called the organizing center (OC), located directly underneath the stem cells. Stem cell maintenance depends in part on a negative feedback loop between the OC and stem cells involving expression of WUSCHEL (WUS) and CLAVATA3 (CLV3).

WUS is expressed in the OC and encodes a homeodomain protein that functions non-cell-autonomously to maintain the stem cells overlying the OC in an undifferentiated state (Mayer et al., 1998). An increase in WUS expression results in an increase in stem cell proliferation. CLV3 is expressed exclusively in stem cells and encodes a small peptide that functions as a ligand of the CLV1/2 receptor kinase complex, which negatively regulates WUS expression and thus delimits the size of the OC (Brand et al., 2000; Schoof et al., 2000). CLV3 also acts non-cell-autonomously. An increase in the number of stem cells leads to an increase in the amount of CLV3 released from stem cells and a corresponding increase in the activity of the CLV1 receptor in these cells. This in turn leads to a decrease in WUS expression and a corresponding decrease in stem cell proliferation (reviewed in Clark, 2001; Nakajima and Benfey, 2002). Lenhard and Laux (2003) showed that while CLV3 restricts the size of the OC by causing inhibition of WUS expression in stem cells and their immediate neighbors, a stable OC (marked by continued WUS expression) is maintained at slightly farther remove underneath the stem cells because CLV3 movement is restricted to just a few cells adjacent to stem cells by its binding to CLV1; thus, it is prevented from entering the underlying OC.

Only a few details of the complex signaling network that influence the WUS-CLV3 feedback loop are understood. Until recently, there has been little information on the downstream targets of WUS that influence meristem size and function. Leibfried et al. (2005) recently reported that WUS directly represses transcription of several two-component ARABIDOPSIS RESPONSE REGULATOR (ARR) genes (ARR5, ARR6, ARR7, and ARR15) in the SAM. These and other type-A ARR genes are rapidly induced by cytokinin, which, together with auxin, exerts strong influence over cell proliferation and differentiation of roots and shoots. WUS repression of ARR genes might be a key factor in maintenance of the SAM. In floral meristems, Lohmann et al. (2001) found that WUS initially activates AGAMOUS (AG) by binding directly to the AG promoter. AG encodes a MADS domain protein that functions to limit proliferation of stem cells in the floral meristem and to specify stamen and carpel identity (Bowman et al., 1989, 1991; Yanofsky et al., 1990). WUS binding to AG was also found to be enhanced by LEAFY, which is strongly expressed in the floral meristem but not in the SAM (Blázquez et al., 1997). Later in flower development, AG in turn acts as a repressor of WUS, thereby terminating the stem cell population in the flower (Lenhard et al., 2001; Lohmann et al., 2001).

WUS appears to fulfill a complementary and independent function in the SAM to the homeodomain KNOX gene SHOOT MERISTEMLESS (STM), another key suppressor of differentiation that is expressed throughout the SAM (Lenhard et al., 2002). STM acts to suppress differentiation by repressing transcription of ASYMMETRIC LEAVES1, which encodes a MYB domain transcription factor that in turn is a negative regulator of the KNOX genes KNAT1 and KNAT2 (Byrne et al., 2000). These KNOX genes are downregulated at sites of organ primordia and thus are implicated with preventing differentiation or promoting cell division in the SAM (Jackson et al., 1994). This complex negative regulatory pathway presumably allows for fine control of meristem maintenance and the formation of organ primordia at appropriate points along the SAM periphery.

In this issue of The Plant Cell, Würschum et al. (pages 295–307) report that Arabidopsis APETALA2 (AP2), previously known for its role in floral patterning and seed development, also functions in stem cell maintenance in the SAM by influencing the WUS-CLV3 feedback loop. The ap2 mutant of Arabidopsis was isolated based on its defects in flower development indicative of a role in specifying floral organ identity (Koornneef et al., 1983; Komaki et al., 1988). Subsequent studies showed that it functions in the establishment of the floral meristem, in regulation of floral homeotic gene expression (Bowman et al., 1989, 1991), and in seed development (Jofuku et al., 1994, 2005; Ohto et al., 2005). Unlike other floral organ specification genes, AP2 shows a broad expression domain throughout the floral meristem and in leaves and stems, leading Jofuku et al. (1994) to conclude that AP2 may play a general role in regulation of development in Arabidopsis. The results of Würschum et al. provide additional support for this prescient conclusion.

Würschum et al. isolated the mutant l28, which displays a semidominant phenotype of premature termination of the shoot meristem and differentiation of stem cells (see figure). They used allele competition experiments together with positional cloning to show that the l28 mutation likely is a dominant-negative allele of AP2. The l28 mutation introduced a single base pair change in the AP2 coding sequence that changed a Glu residue to Lys in one of two AP2 domains thought to be involved in DNA binding. Since previous work had not indicated a role for AP2 in shoot meristem maintenance, the authors were interested to know whether the l28 mutation points to a novel function for AP2 or whether it creates an abnormal protein that interferes with processes in the shoot meristem not normally influenced by wild-type AP2. To address this question, they used heterozygous l28 mutant plants, which contain both the wild-type AP2 allele and the dominant-negative l28 allele, and sought to determine if the wild-type and mutant proteins competed for the same targets in vivo. This was accomplished by reducing wild-type AP2 activity in the l28 heterozygotes in a dose-dependent manner by crossing the l28 mutants with homozygous mutants of ap2-1 (a weak loss-of-function allele) and ap2-2 (a putative null allele), generating a dosage series of AP2 activity.

View larger version (124K): [in this window] [in a new window]   l28 Mutant Phenotype. Wild-type Arabidopsis seedlings (left) have initiated rosette leaves from the SAM, whereas l28 mutants (right) have terminated the shoot apical meristem prematurely. Bars = 5 mm.

The authors found that AP2 is strongly expressed in all tissues from early embryo stages on, but expression decreases in cells that begin to undergo differentiation, consistent with a role in meristem maintenance. They further found that shoot meristem size was reduced in the l28 mutant relative to the wild type, and more specifically, the expression of both WUS and CLV3 was severely repressed or abolished by the l28 mutation. Because AG has been found to interact with WUS (WUS activating AG expression early and AG repressing WUS late in flower development; Lenhard et al., 2001; Lohmann et al., 2001) and is itself repressed by AP2 (Drews et al., 1991), Würschum et al. next asked whether the l28 mutation affects WUS via ectopic expression of AG. However, the segregating progeny of plants heterozygous for l28 and ag-1 showed no difference in shoot meristem termination relative to homozygous l28 mutants, suggesting that AP2 functions independently of AG in maintenance of the stem cell niche.

Like the l28 mutant seedlings, seedlings carrying the putative null wus-1 allele display a flat apex of differentiated cells in place of a SAM and lack CLV3 expression. Crossing heterozygous l28 mutants with heterozygous wus-1 mutants further suggested that both the l28 and wus-1 mutations disrupt the stem cell niche of the SAM in a similar manner. With regard to CLV3, the authors found that clv3 mutations rescued shoot meristem development in the l28 mutant background, indicating that termination of the shoot meristem by l28 requires an active CLV3 gene.

This work reveals another important link in the WUS-CLV3 feedback loop regulating the stem cell niche in Arabidopsis. Würschum et al. propose a plausible model in which AP2 (or factors regulated by it) affect stem cell maintenance by negatively regulating the CLV signaling pathway because the l28 phenotype requires an active CLV3 gene. It is also possible that WUS is a target of AP2 or that WUS and AP2 interact with some of the same downstream targets that exert feedback control on CLV signaling. Since WUS expression in the SAM is restricted to the OC, whereas AP2 is expressed throughout the meristem, it may be that WUS acts on a subset of a larger group of AP2-interacting factors or downstream targets. Clearly one of the next steps will be identifying AP2 interacting partners in the SAM.

It might be of interest to investigate whether AP2 is linked to STM activity in the SAM in a previously undiscovered fashion. In this regard, Kirch et al. (2003) identified DORNRÖSCHEN/ENHANCER OF SHOOT REGENERATION1 (DRN/ESR1), which encodes an AP2/ERF domain protein that appears to influence the expression of STM, WUS, and CLV3. Banno et al. (2001) earlier characterized this gene as ESR1 because overexpression of the gene conferred cytokinin-independent shoot formation in Arabidopsis root explants. Kirch et al. (2003) identified DRN/ESR1 via activation tagging: drn/esr1 gain-of-function mutants prematurely arrested shoot meristem activity and formed radialized lateral organs. DRN/ESR1 was found to be expressed in a subdomain of meristem cells and lateral organ primordia (anlagen), and no phenotype was associated with loss-of-function mutations, suggesting that DRN/ESR1 is a redundant component involved in stem cell maintenance in the Arabidopsis SAM.

	REFERENCES

TOP REFERENCES

 
Banno, H., Ikeda, Y., Niu, Q.-W., and Chua, N.-H. (2001). Overexpression of Arabidopsis ESR1 induces initiation of shoot regeneration. Plant Cell 13, 2609–2618.[Abstract/Free Full Text]

Blázquez, M.E., Soowal, L.N., Lee, I., and Weigel, D. (1997). LEAFY expression and flower initiation in Arabidopsis. Development 124, 3835–3844.[Abstract]

Bowman, J.L., Smyth, D.R., and Meyerowitz, E.M. (1989). Genes directing flower development in Arabidopsis. Plant Cell 1, 37–52.[Abstract/Free Full Text]

Bowman, J.L., Smyth, D.R., and Meyerowitz, E.M. (1991). Genetic interactions among floral homeotic genes of Arabidopsis. Development 112, 1–20.[Abstract]

Brand, U., Fletcher, J.C., Hobe, M., Meyerowitz, E.M., and Simon, R. (2000). Dependence of stem cell fate in Arabidopsis on a feedback loop regulated by CLV3 activity. Science 289, 617–619.[Abstract/Free Full Text]

Byrne, M.E., Barley, R., Curtis, M., Arroyo, J.M., Dunham, A., Hudson, A., and Martienssen, R.A. (2000). Asymmetric leaves1 mediates leaf patterning and stem cell function in Arabidopsis. Nature 408, 967–971.[CrossRef][Medline]

Clark, S.E. (2001). Cell signalling at the shoot meristem. Nat. Rev. Mol. Cell Biol. 2, 276–284.[CrossRef][ISI][Medline]

Drews, G.N., Bowman, J.L., and Meyerowitz, E.M. (1991). Negative regulation of the Arabidopsis homeotic gene AGAMOUS by the APETALA2 product. Cell 65, 991–1002.[CrossRef][ISI][Medline]

Jackson, D., Veit, B., and Hake, S. (1994). Expression of maize KNOTTED1 related homeobox genes in the shoot apical meristem predicts patterns of morphogenesis in the vegetative shoot. Development 120, 405–413.[Abstract]

Jofuku, K.D., Boer, B.G.W.d., Montagu, M.V., and Okamuro, J.K. (1994). Control of Arabidopsis flower and seed development by the homeotic gene APETALA2. Plant Cell 6, 1211–1225.[Abstract]

Jofuku, K.D., Omidyar, P.K., Gee, Z., and Okamuro, J.K. (2005). Control of seed mass and seed yield by the floral homeotic gene APETALA2. Proc. Natl. Acad. Sci. USA 102, 3117–3122.[Abstract/Free Full Text]

Kirch, T., Simon, R., Grünewald, M., and Werr, W. (2003). The DORNRÖSCHEN/ENHANCER OF SHOOT REGENERATION1 gene of Arabidopsis acts in the control of meristem cell fate and lateral organ development. Plant Cell 15, 694–705.[Abstract/Free Full Text]

Komaki, M.K., Okada, K., Nishino, E., and Shimura, Y. (1988). Isolation and characterization of nove1 mutants of Arabidopsis thaliana defective in flower development. Development 104, 195–203.[Abstract]

Koornneef, M., Vaneden, J., Hanhart, C.J., Stam, P., Braakmsa, F.J., and Feenstra, W.J. (1983). Linkage map of Arabidopsis thaliana. J. Hered. 74, 265–272.[Abstract/Free Full Text]

Leibfried, A., To, J.P.C., Busch, W., Stehling, S., Kehle, A., Demar, M., Kieber, J.J., and Lohmann, J.U. (2005). WUSCHEL controls meristem function by direct regulation of cytokinin-inducible response regulators. Nature 438, 1172–1175.[CrossRef][Medline]

Lenhard, M., Bohnert, A., Jürgens, G., and Laux, T. (2001). Termination of stem cell maintenance in Arabidopsis floral meristems by interactions between WUSCHEL and AGAMOUS. Cell 105, 805–814.[CrossRef][ISI][Medline]

Lenhard, M., Jürgens, G., and Laux, T. (2002). The WUSCHEL and SHOOT MERISTEMLESS genes fulfill complementary roles in Arabidopsis shoot meristem regulation. Development 129, 3195–3206.[Abstract/Free Full Text]

Lenhard, M., and Laux, T. (2003). Stem cell homeostasis in the Arabidopsis shoot meristem is regulated by intercellular movement of CLAVATA3 and its sequestration by CLAVATA1. Development 130, 3163–3173.[Abstract/Free Full Text]

Lohmann, J., Huong, R., Hobe, M., Busch, M., Parcy, F., Simon, R., and Weigel, D. (2001). A molecular link between stem cell regulation and floral patterning in Arabidopsis. Cell 105, 793–803.[CrossRef][ISI][Medline]

Mayer, K.F.X., Schoof, H., Haecker, A., Lenhard, M., Jürgens, G., and Laux, T. (1998). Role of WUSCHEL in regulating stem cell fate in the Arabidopsis shoot meristem. Cell 95, 805–815.[CrossRef][ISI][Medline]

Nakajima, K., and Benfey, P.N. (2002). Signaling in and out: Control of cell division and differentiation in the shoot and root. Plant Cell 14 (suppl.), S265–S276.[Free Full Text]

Ohto, M., Fischer, R.L., Goldberg, R.B., Nakamura, K., and Harada, J.J. (2005). Control of seed mass by APETALA2. Proc. Natl. Acad. Sci. USA 102, 3123–3128.[Abstract/Free Full Text]

Schoof, H., Lenhard, M., Haecker, A., Mayer, K.F.X., Jürgens, G., and Laux, T. (2000). The stem cell population of Arabidopsis shoot meristems is maintained by a regulatory loop between the CLAVATA and WUSCHEL genes. Cell 100, 635–644.[CrossRef][ISI][Medline]

Würschum, T., Groß-Hardt, R., and Laux, T. (2006). APETALA2 regulates the stem cell niche in the Arabidopsis shoot meristem. Plant Cell 18, 295–307.[Abstract/Free Full Text]

Yanofsky, M.F., Ma, H., Bowman, J.L., Drews, G.N., Feldmann, K.A., and Meyerowitz, E.M. (1990). The protein encoded by the Arabidopsis homeotic gene AGAMOUS resembles transcription factors. Nature 346, 35–39.[CrossRef][Medline]

Related articles in Plant Cell:

APETALA2 Regulates the Stem Cell Niche in the Arabidopsis Shoot Meristem

Tobias Würschum, Rita Groß-Hardt, and Thomas Laux
Plant Cell 2006 18: 295-307. [Abstract] [Full Text]  

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Related articles in Plant Cell Similar articles in this journal Similar articles in ISI Web of Science Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Eckardt, N. A. Search for Related Content PubMed Articles by Eckardt, N. A. Agricola Articles by Eckardt, N. A.