Skip to main content
NCBI home page
Elsevier Sponsored Documents logoLink to Elsevier Sponsored Documents
. 2014 Aug 18;24(16):1939–1944. doi: 10.1016/j.cub.2014.07.019

WOX5 Suppresses CYCLIN D Activity to Establish Quiescence at the Center of the Root Stem Cell Niche

Celine Forzani 1, Ernst Aichinger 2, Emily Sornay 1, Viola Willemsen 3, Thomas Laux 2, Walter Dewitte 1,∗, James AH Murray 1,∗∗
PMCID: PMC4148176  PMID: 25127220

Summary

In Arabidopsis, stem cells maintain the provision of new cells for root growth. They surround a group of slowly dividing cells named the quiescent center (QC), and, together, they form the stem cell niche (SCN). The QC acts as the signaling center of the SCN, repressing differentiation of the surrounding stem cells [1] and providing a pool of cells able to replace damaged stem cells [2, 3]. Maintenance of the stem cells depends on the transcription factor WUSCHEL-RELATED HOMEOBOX 5 (WOX5), which is specifically expressed in the QC [4]. However, the molecular mechanisms by which WOX5 promotes stem cell fate and whether WOX5 regulates proliferation of the QC are unknown. Here, we reveal a new role for WOX5 in restraining cell division in the cells of the QC, thereby establishing quiescence. In contrast, WOX5 and CYCD3;3/CYCD1;1 both promote cell proliferation in the nascent columella. The additional QC divisions occurring in wox5 mutants are suppressed in mutant combinations with the D type cyclins CYCD3;3 and CYCD1;1. Moreover, ectopic expression of CYCD3;3 in the QC is sufficient to induce cell division in the QC. WOX5 thus suppresses QC divisions that are otherwise promoted by CYCD3;3 and CYCD1;1, in part by interacting with the CYCD3;3 promoter to repress CYCD3;3 expression in the QC. Therefore, we propose a specific role for WOX5 in initiating and maintaining quiescence of the QC by excluding CYCD activity from the QC.

Graphical Abstract

graphic file with name fx1.jpg

Open in a new tab

Highlights

  • •

    WOX5 prevents divisions at the root stem cell niche center to initiate quiescence

  • •

    WOX5 suppresses CYCD expression in the quiescent center to restrict cell divisions

  • •

    WOX5 binds to the CYCD3;3 promoter

  • •

    CYCD3;3 and CYCD1;1 stimulate division during formation of the columella

Forzani et al. reveal a critical role for the WOX5 transcription factor in initiating and maintaining quiescence at the center of the root stem cell niche in Arabidopsis. They demonstrate that WOX5 locally suppresses cell division by preventing expression of the D type cyclin CYCD3;3 in the quiescent center.

Results and Discussion

WOX5 Stimulates Columella Development and Initiates Quiescence in the Embryonic QC

The root apical meristem (RAM) is initiated during embryogenesis when the uppermost suspensor cell or hypophysis divides asymmetrically to produce an upper lens-shaped cell, the progenitor of the quiescent center (QC), and a larger basal cell that gives rise to the columella stem cells (CSCs) and columella [5]. The QC maintains a low frequency of division, which reduces vulnerability to DNA damage [2, 3], and also specifies adjacent cells to become the stem cells that actively divide to renew themselves [6, 7]. How this quiescence is initiated and maintained in the midst of proliferating stem cells is unknown. Because WUSCHEL-RELATED HOMEOBOX 5 (WOX5) expression is first detected in the hypophysis and remains specifically expressed in the QC during both embryogenesis and postgermination growth [4, 8], we tested whether WOX5 regulates cell division in the descendants of the hypophysis.

We used mature embryos as a “read out” of embryonic development in which cell numbers and patterns are easily studied. WOX5 is required postembryonically to maintain CSCs in the RAM [4], so we reasoned it might also be active during embryogenesis when the columella is forming. Therefore, we counted the number of columella cells (CCs) distal to the QC and the number of columella layers in two loss-of-function alleles of WOX5 (Figures 1A–1C). The number of columella layers was reduced in wox5 embryos, with 80% showing three layers of CCs instead of four in wild-type (WT) Col-0 (Figures 1E and 1F). Occasionally, enlarged CCs spanning two layers were observed in 92% of wox5 mutant roots (Figures 1B and 1C). Introducing a construct carrying the WOX5 locus into wox5-1 mutants completely restored the number of columella layers (Figures 1D–1F; Figures S1B and S1C available online), confirming that the observed defects were caused by the wox5-1 mutation.

Figure 1.

Figure 1

Open in a new tab

During Embryogenesis, WOX5 Suppresses Cell Division in the QC and Induces Divisions in the Columella

(A–D) Cell walls in mature embryos were visualized by propidium iodide staining. wox5 mutants display transverse divisions in the QC, as indicated by the yellow arrow, and a reduced number of CCs compared to WT Col-0. Scale bars represent 10 μm.

(A) Mature embryo with the different cell types color coded: QC in blue; columella in purple.

(D) The WOX5 genomic DNA was expressed from the WOX5 promoter in wox5-1.

(E) Quantification of the number of cells in the embryonic root. The number of cells in the columella was counted according to the scheme in (A).

(F) Quantification of the number of layers in the columella. The maximum number of layers was counted (see yellow dots in A).

(G) Quantification of the number of transverse divisions occurring in the QC of mature embryos or 3-day-old roots.

(E–G) Numbers of embryos examined: WT Col-0, n = 73; wox5-1, n = 91; wox5-3, n = 85; pWOX5:WOX5, n = 45. Thirty seedlings were counted for the WT Col-0 or for the mutant wox5-1. Error bars show SEs (E and G). Student’s t test, ∗p < 0. 00001.

QC, quiescent center; DAG, days after germination. See also Figure S1.

This led us to conclude that WOX5 functions in embryogenesis to ensure the correct number of cells in the columella. Because WOX5 is expressed specifically in the embryonic QC, we tested whether loss of WOX5 also affected the cellular organization of the QC and specifically the initiation of quiescence. QC cell divisions were quantified by determining the number of transverse divisions (Figures 1B and 1C). wox5 embryonic roots showed an increase in QC cell divisions, with on average 1.4 divisions occurring per root compared to 0.013 divisions in WT Col-0 (Figure 1G), and this was suppressed in wox5-1 mutants expressing a pWOX5:WOX5 construct (Figures 1D, 1G, and S1A). Furthermore, the increase in QC cell division was maintained after germination in the primary root (Figure 1G).

These results demonstrate that in addition to its previously documented role in maintaining CSCs postgermination [4], WOX5 mediates cell numbers in the nascent columella and cell autonomously restricts cell division in the QC.

CYCD1;1 and CYCD3;3 Control Cell Division during the Morphogenesis of the Columella and Prevent Premature Stem Cell Differentiation

The molecular mechanisms regulating cell division in the developing root pole are currently unknown. The mitotic cell cycle is driven by the action of cyclin-dependent kinase (CDK)/cyclin complexes, and CDKA/cyclin D (CYCD) complexes control the commitment point at the G1/S transition through reversible phosphorylation of the RETINOBLASTOMA-RELATED (RBR) protein [9, 10, 11, 12]. RBR has recently been shown to act cell autonomously in the QC to limit cell divisions [2, 13], and it plays a key role in restraining cell proliferation in root stem cells [14]. However, RBR is encoded by a single gene expressed throughout the root [14]. Because the Arabidopsis genome contains ten CYCD genes with distinct patterns of expression in the root [15, 16], these are more likely candidates to provide tissue-specific regulation of RBR activity. Furthermore, elevating CYCD3;1 levels results in additional CSC layers [14], and CYCDs respond to intrinsic and extrinsic signals [17, 18, 19].

To investigate whether specific CYCDs control cell division in the developing root pole and stem cell niche, we screened mature embryos of single loss-of-function alleles of the various CYCDs for defects in cell division in the columella (Figure S2A). Only cycd3;3 mutants showed a reduced number of CCs, with 25% of cycd3;3 embryos having three columella layers instead of four (Figures 2A, 2B, and 2M). Because the Arabidopsis CYCD3 subgroup has three members (CYCD3;1, CYCD3;2, CYCD3;3) [18], different combinations of the cycd3 mutant alleles were assessed for enhanced defects with respect to cell division in the columella (Figure S2A). cycd3;1-3 embryos lacking all CYCD3 function were very similar to cycd3;3 embryos, indicating that from the CYCD3 subgroup, only CYCD3;3 is essential for normal cell division activity in the embryonic columella.

Figure 2.

Figure 2

Open in a new tab

Genetic Interaction between CYCD1;1/CYCD3;3 and WOX5

(A–H) Mature embryos were visualized by propidium iodide staining. Yellow arrows indicate transverse divisions occurring in the QC.

(A–D) cycd;3;3 and cycd1;1 cycd3;3 mutants show a reduced number of CCs in the embryonic root compared to WT Col-0.

(E–H) The transverse divisions occurring in the wox5-1 QC are absent from the wox5 cycd1;1 cycd3;3 QC.

(I) pCYCD1;1:GUS-GFP reporter line shows GFP expression in the embryonic QC. pCYCD3;3:GUS-GFP reporter line shows GFP expression in the embryonic root pole.

(J) Left: diagram illustrating the Arabidopsis root with the QC (colored in red), one layer of CSCs, and four layers of CCs. Right: percentage of roots with zero, one, or two layers of CSCs in 5-day-old roots. Error bars show SD from three biological repeats. Between 30 and 40 roots were examined for every genotype per experiment.

(K) QC46 expression in WT Col-0 and cycd1;1 cycd3;3 of 5-day-old seedlings. The enhancer trap line QC46 expresses β-glucuronidase in the QC (GUS activity is indicated by the blue arrow). Starch granules in amyloplasts were stained dark blue by lugol.

(L) QC46 expression in the QC of mature embryos.

(M) Quantification of the number of layers in the columella. The maximum number of layers was counted (see yellow dots in A).

(N) Quantification of the number of transverse divisions occurring in the QC of mature embryos (N) or 3-day-old roots (O).

(M and N) Numbers of embryos examined: Col-0, n = 108; cycd3;3, n = 101; cycd1;1, n = 102; cycd1;1cycd3;3, n = 119; wox5-1, n = 107; wox5 cycd1;1, n = 118; wox5 cycd3;3, n = 109; wox5 cycd1;1 cycd3;3, n = 116.

(O) Twenty-five seedlings were counted for each genotype.

Error bars show SEs (N and O). Student’s t test, ∗p < 0.001, ∗∗p < 0.01. The asterisk (∗) in (O) indicates a statistical difference between wox5-1 and the different mutant combinations. Scale bars of (A)–(I) and (L) represent 10 μm. QC, quiescent center; DAG, days after germination. See also Figure S2.

Because redundancy with other CYCD genes might be responsible for the relatively weak cycd3;3 mutant phenotype, we examined the tissue specificity of their expression [16] to identify other potential cofunctioning candidates. Monitoring the GFP fluorescence from CYCD promoters driving a GUS-GFP reporter construct showed that only CYCD1;1 displayed specific expression in the embryonic QC, persisting from heart to torpedo stage (Figure 2I) [16]. In contrast, CYCD3;3 expression was not observed in the QC but was apparent in the root cap from heart until torpedo stage (Figure 2I). Given that WOX5 acts from the QC to maintain CSCs, we tested whether CYCD1;1 might similarly influence the divisions in the columella from its site of expression in the QC. The cycd1;1 mutants showed no detectable phenotype, but, in combination with cycd3;3, they did enhance the cycd3;3 phenotype (Figures 2C and 2D). One hundred percent of cycd1;1 cycd3;3 embryos displayed three columella layers instead of four (Figure 2M), and occasionally, enlarged CCs spanning two layers were observed in 56% of cycd1;1 cycd3;3 mutant roots.

After germination, CCs are generated by the dividing CSCs at the distal side of the QC. Differentiated CCs accumulate amyloplasts, starch-accumulating plastids, which are absent from CSCs. WOX5 is required to maintain the identity of the CSCs, which differentiate in wox5 mutants [4].

Given that the embryonic columella phenotype of cycd1;1 cycd3;3 mutants is similar to that of wox5, we assessed whether CYCD1;1 and CYCD3;3 are required to maintain CSCs in the primary root. One or two layers of CSCs are detected in 5-day-old WT roots (Figure 2J). However, in 35% of cycd1;1 cycd3;3 roots, amyloplasts accumulated in CSCs, indicating a loss of CSC identity and premature differentiation into CCs (Figure 2J). cycd3;3 mutant roots also showed early differentiation, albeit to a lesser extent than cycd1;1 cycd3;3.

Because QC function is essential to repress CSC differentiation, we tested whether QC identity was modified in cycd1;1 cycd3;3 roots. Three different QC-specific molecular markers, QC25 (Figure S2B), QC46 (Figure 2K) [20], and WOX5:ER-GFP (Figure S2B) [21], were normally expressed in the QC of cycd1;1 cycd3;3 mutant roots, suggesting that the QC identity is unaffected.

To confirm that the phenotypes observed in cycd1;1 cycd3;3 mutants were caused by loss-of-function of these cyclins, we examined a second loss-of-function allele of CYCD1;1, cycd1;1-2, in combination with cycd3;3. The cycd1;1-2 cycd3;3 double mutant recapitulated the original cycd1;1 cycd3;3 phenotype (Figures S2C and S2D). In the absence of a second loss-of-function allele of CYCD3;3, we complemented cycd3;3 and cycd1;1 cycd3;3 double mutants with a construct encoding vYFP fused to a CYCD3;3 genomic fragment under the control of the CYCD3;3 promoter. This efficiently rescued both cycd3;3 and cycd1;1 cycd3;3 abnormalities (Figure S2E).

Taken together, we conclude that CYCD1;1 and CYCD3;3 stimulate cell division during the formation of the columella and prevent CSC differentiation to maintain the CSC niche postgermination.

WOX5 Acts through CYCD1;1 and CYCD3;3 to Control Cell Divisions in the QC

To assess whether WOX5 might act through CYCD1;1 and CYCD3;3 to regulate cell division, we tested the genetic interaction of wox5-1 with the cycd1;1 cycd3;3 mutant. The additional QC cell divisions observed in 89% of wox5-1 embryonic roots were almost totally suppressed in wox5 cycd1;1 cycd3;3 (8%; Figures 2A–2H and 2N). This phenotype was already apparent in wox5 cycd3;3 double mutant, in which only 45% of embryos showed divisions in the QC. Hence, both CYCD1;1 and CYCD3;3 contribute to the extra transverse divisions observed in the wox5-1 embryonic QC. The identity of the QC was confirmed through expression of the QC-specific marker QC46 [20]. QC46 was normally expressed in the mature embryonic QC of wox5-1 cycd1;1 cycd3;3 as it was in wox5-1 or cycd1;1 cycd3;3 roots (Figure 2L), indicating that QC development was unaffected.

We then tested whether CYCD1;1 and CYCD3;3 fulfil the same role postembryonically, being rate limiting for QC cell division in wox5-1 mutant roots. Indeed, the additional QC cell divisions present in wox5-1 mutants were reduced by 45% in wox5 cycd3;3 and by 55% in wox5 cycd1;1 cycd3;3 mutants (Figure 2O). However, we note that additional QC cell divisions were also seen in cycd1;1 cycd3;3 roots. This would be consistent with the premature differentiation of the CSCs in the cycd1;1 cycd3;3 roots (Figure 2J; Table S1), triggering additional cell divisions of the QC to replenish the CSC pool.

Taken together, these results suggest that CYCD1;1 and CYCD3;3 drive the abnormal divisions in the wox5-1 QC.

In contrast, in the developing columella, WOX5 seems to act, in part, independently from CYCD3;3 and CYCD1;1. A further reduction in the number of CCs was observed in the triple wox5-1 cycd1;1 cycd3;3 mutant compared to either wox5-1 or cycd1;1 cycd3;3. Fifty-seven percent of wox5-1 cycd1;1 cycd3;3 embryos had only two columella layers compared to 4% in wox5-1 and 0% in cycd1;1 cycd3;3 (Figures 2M and S2F). These results suggest that WOX5 may partially act through CYCD1;1/CYCD3;3 and also through additional unknown targets to control cell division in the embryonic columella.

Ectopic QC Expression of CYCD3;3 Induces QC Cell Division

To further assess whether CYCD regulates cell division in the QC, we tested whether ectopic expression of CYCD3;3 in the QC would be sufficient to induce cell division. Both the vYFP-CYCD3;3 and the vYFP-CYCD1;1 fusion proteins expressed under the control of the WOX5 promoter triggered transverse divisions in the QC. Both proteins were consistently detected in the nucleus of the QC (Figure S3B), and vYFP-CYCD3;3 triggered QC divisions in 100% of embryos and vYFP-CYCD1;1 in 9% (Figures 3A–3D and S3A).

Figure 3.

Figure 3

Open in a new tab

Ectopic CYCD Expression in the QC Is Sufficient to Induce QC Cell Division

(A–C) Mature embryos were visualized by propidium iodide staining. Yellow arrows indicate transverse divisions occurring in the QC of pWOX5:vYFP-CYCD1;1 and pWOX5:vYFP-CYCD3;3 expressed in WT Col-0. The CYCD1;1 or the CYCD3;3 genomic DNA was expressed from the WOX5 promoter.

(D) Quantification of the number of transverse divisions occurring in the QC of mature embryos. Numbers of embryos examined: WT Col-0, n = 40; pWOX5:vYFP-CYCD1;1, n = 33; pWOX5:vYFP-CYCD3;3, n = 34.

(E) QC46 and QC25 expression in the QC of mature embryos of WT Col-0 (reduced in Col-0 expressing either pWOX5:vYFP-CYCD1;1 or pWOX5:vYFP-CYCD3;3). F1 populations were used with QC46 and QC25, being hemizygous in all the genotypes.

Scale bars of (A)–(C) and (E) represent 10 μm. See also Figure S3.

To analyze whether these additional divisions influenced QC identity, we examined the expression of the QC-specific markers, QC46 and QC25, in the embryonic root (Figure 3E) and pWOX5:ER-GFP in the QC of 5-day-old roots (Figure S3C). Although activity of these QC-specific markers is apparent in WT Col-0 lines expressing pWOX5:vYFP-CYCD1;1 or pWOX5:vYFP-CYCD3;3, all showed reduced expression, suggesting a potential partial loss of QC identity (Figures 3E and S3C).

These results suggest that restraining of CYCD activity in the QC is required to maintain both quiescence and the correct cellular identity in the QC.

CYCD3;3 Expression Is Suppressed by WOX5 in the QC

To establish whether WOX5 acts to exclude CYCD expression from the QC, thereby maintaining its quiescence, we compared CYCD1;1 or CYCD3;3 expression in wox5-1 and WT mutant roots by using a GUS-GFP fusion protein expressed under the control of the CYCD promoter. Differences were seen only for CYCD3;3 expression, which was detected in the QC of wox5-1 seedlings but was almost absent from WT QC (Figure 4A). In WT roots, CYCD3;3 was also expressed in the distal columella layers, the lateral root cap, the epidermal stem cells, and the stele, but this pattern was unaltered in wox5-1 mutant roots. This CYCD3;3 promoter is functional in planta because vYFP-CYCD3;3 expressed under its control complements the cycd1;1 cycd3;3 mutant phenotype (Figure S2E). These results were reproducible with three independent pCYCD3;3:GUS-GFP lines crossed to wox5-1 mutants and suggest that WOX5 is required to exclude CYCD3;3 expression from the QC.

Figure 4.

Figure 4

Open in a new tab

WOX5 Restricts CYCD3;3 Activity from the QC and Binds to Its Promoter

(A) pCYCD3;3:GUS-GFP expression in wox5-1 roots expands into the QC compared to WT Col-0 roots. Cell walls of 5-day-old roots were stained with propidium iodide. Scale bars represent 10 μm.

(B and C) ChIP experiments were performed with root samples from WT Col-0 plants expressing a p35S:WOX5-GR construct and wox5-1 mutant plants. An anti-WOX5 antibody was used to immunoprecipitate WOX5, followed by qPCR of the CYCD3;3 promoter and gene as illustrated in (B) and of the CYCD3;1 and CYCD6;1 promoter regions. The fold enrichment was calculated relative to the input and signal at the eukaryotic initiation factor (eIF4) gene. Error bars show SE from three technical replicates. The ChIP experiment was repeated three times, with similar results each time.

To assess whether this downregulation is direct, we tested whether WOX5 binds the CYCD3;3 promoter by using chromatin immunoprecipitation (ChIP)-quantitative PCR (qPCR) assays. Two of five regions tested from the CYCD3;3 promoter, located 700 base pairs upstream of the transcriptional start site (TSS), and a fragment spanning the TSS showed 6.5-fold and 3.5-fold enrichment, respectively, compared to adjacent promoter regions (Figure 4B) and to wox5-1 mutant controls (Figure 4C). In addition, no enrichment was seen for the CYCD3;1 and the CYCD6;1 promoter fragments, suggesting that WOX5 binding is not a conserved feature among CYCD promoters.

Taken together, we conclude that WOX5 binds the CYCD3;3 promoter directly or indirectly, and it negatively regulates its expression in the QC.

Conclusions

Here, we demonstrate a new role for the transcription factor WOX5 in establishing and maintaining quiescence of the QC by regulating CYCD3;3 activity. This establishes a direct link between core cell-cycle components and transcription factors in organizing the root stem cell niche. We observe that CYCD1;1 and CYCD3;3 stimulate cell division in the descendants of the hypophysis to form the embryonic columella. During embryogenesis, local action of WOX5 initiates quiescence in the QC, whose maintenance requires the continuing presence of WOX5. WOX5 acts in the QC through local suppression of CYCD3;3 expression, and this function is needed to restrict cell division. Furthermore, an appropriate level of CYCD3;3 and CYCD1;1 is required for normal QC development. We conclude that tight control of CYCD activity by WOX5 is essential for QC quiescence and for the normal development of the QC. In contrast, in the nascent columella, WOX5 induces cell proliferation by acting on different targets and not primarily through CYCD3;3 and CYCD1;1.

Hormone [22, 23, 24, 25] and peptide [2] signaling pathways are known to regulate cell division in the QC, and recent advances have shown that a low proliferation rate of the QC is essential to maintain plant growth during stressful conditions [2, 3]. It remains to be shown how WOX5 and CYCD presumably integrate these different signaling pathways to regulate cell division in the QC and maintain a functional stem cell niche during normal or stressed conditions.

Acknowledgments

We thank Angela Marchbank and Joanne Kilby for excellent technical support. This project was supported by grants BB/E024858, BB/J009199/1, and BB/L009358/1 from the Biotechnology and Biological Sciences Research Council (BBSRC) to J.A.H.M. and W.D., by the ERA-NET in Plant Genomics (ERA-PG; project “Plant Stem Cell Network”; to J.A.H.M., W.D., and T.L.), by the EU-cofunded INTERREG IV Upper Rhine Project 692 A17 (to T.L.), by the Deutsche Forschungsgemeinschaft (SFB592), by the Excellence Initiative of the German federal and state governments (FRIAS; to T.L.), and by an EU Marie Curie Fellowship to E.A.

Published: August 7, 2014

Footnotes

This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/3.0/).

Supplemental Information includes Supplemental Experimental Procedures, three figures, and one table and can be found with this article online at http://dx.doi.org/10.1016/j.cub.2014.07.019.

Contributor Information

Walter Dewitte, Email: dewittew@cardiff.ac.uk.

James A.H. Murray, Email: murrayja1@cardiff.ac.uk.

Supplemental Information

Document S1. Supplemental Experimental Procedures, Figures S1–S3, and Table S1
mmc1.pdf (3.2MB, pdf)
Document S2. Article plus Supplemental Information
mmc2.pdf (4.8MB, pdf)

References

  • 1.van den Berg C., Willemsen V., Hendriks G., Weisbeek P., Scheres B. Short-range control of cell differentiation in the Arabidopsis root meristem. Nature. 1997;390:287–289. doi: 10.1038/36856. [DOI] [PubMed] [Google Scholar]
  • 2.Cruz-Ramírez A., Díaz-Triviño S., Wachsman G., Du Y., Arteága-Vázquez M., Zhang H., Benjamins R., Blilou I., Neef A.B., Chandler V., Scheres B. A SCARECROW-RETINOBLASTOMA protein network controls protective quiescence in the Arabidopsis root stem cell organizer. PLoS Biol. 2013;11:e1001724. doi: 10.1371/journal.pbio.1001724. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Heyman J., Cools T., Vandenbussche F., Heyndrickx K.S., Van Leene J., Vercauteren I., Vanderauwera S., Vandepoele K., De Jaeger G., Van Der Straeten D., De Veylder L. ERF115 controls root quiescent center cell division and stem cell replenishment. Science. 2013;342:860–863. doi: 10.1126/science.1240667. [DOI] [PubMed] [Google Scholar]
  • 4.Sarkar A.K., Luijten M., Miyashima S., Lenhard M., Hashimoto T., Nakajima K., Scheres B., Heidstra R., Laux T. Conserved factors regulate signalling in Arabidopsis thaliana shoot and root stem cell organizers. Nature. 2007;446:811–814. doi: 10.1038/nature05703. [DOI] [PubMed] [Google Scholar]
  • 5.Dolan L., Janmaat K., Willemsen V., Linstead P., Poethig S., Roberts K., Scheres B. Cellular organisation of the Arabidopsis thaliana root. Development. 1993;119:71–84. doi: 10.1242/dev.119.1.71. [DOI] [PubMed] [Google Scholar]
  • 6.Aichinger E., Kornet N., Friedrich T., Laux T. Plant stem cell niches. Annu. Rev. Plant Biol. 2012;63:615–636. doi: 10.1146/annurev-arplant-042811-105555. [DOI] [PubMed] [Google Scholar]
  • 7.Lau S., Slane D., Herud O., Kong J., Jürgens G. Early embryogenesis in flowering plants: setting up the basic body pattern. Annu. Rev. Plant Biol. 2012;63:483–506. doi: 10.1146/annurev-arplant-042811-105507. [DOI] [PubMed] [Google Scholar]
  • 8.Haecker A., Gross-Hardt R., Geiges B., Sarkar A., Breuninger H., Herrmann M., Laux T. Expression dynamics of WOX genes mark cell fate decisions during early embryonic patterning in Arabidopsis thaliana. Development. 2004;131:657–668. doi: 10.1242/dev.00963. [DOI] [PubMed] [Google Scholar]
  • 9.Nieuwland J., Scofield S., Murray J.A. Control of division and differentiation of plant stem cells and their derivatives. Semin. Cell Dev. Biol. 2009;20:1134–1142. doi: 10.1016/j.semcdb.2009.09.011. [DOI] [PubMed] [Google Scholar]
  • 10.Boniotti M.B., Gutierrez C. A cell-cycle-regulated kinase activity phosphorylates plant retinoblastoma protein and contains, in Arabidopsis, a CDKA/cyclin D complex. Plant J. 2001;28:341–350. doi: 10.1046/j.1365-313x.2001.01160.x. [DOI] [PubMed] [Google Scholar]
  • 11.Nakagami H., Kawamura K., Sugisaka K., Sekine M., Shinmyo A. Phosphorylation of retinoblastoma-related protein by the cyclin D/cyclin-dependent kinase complex is activated at the G1/S-phase transition in tobacco. Plant Cell. 2002;14:1847–1857. doi: 10.1105/tpc.002550. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Nowack M.K., Harashima H., Dissmeyer N., Zhao X., Bouyer D., Weimer A.K., De Winter F., Yang F., Schnittger A. Genetic framework of cyclin-dependent kinase function in Arabidopsis. Dev. Cell. 2012;22:1030–1040. doi: 10.1016/j.devcel.2012.02.015. [DOI] [PubMed] [Google Scholar]
  • 13.Wachsman G., Heidstra R., Scheres B. Distinct cell-autonomous functions of RETINOBLASTOMA-RELATED in Arabidopsis stem cells revealed by the Brother of Brainbow clonal analysis system. Plant Cell. 2011;23:2581–2591. doi: 10.1105/tpc.111.086199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Wildwater M., Campilho A., Perez-Perez J.M., Heidstra R., Blilou I., Korthout H., Chatterjee J., Mariconti L., Gruissem W., Scheres B. The RETINOBLASTOMA-RELATED gene regulates stem cell maintenance in Arabidopsis roots. Cell. 2005;123:1337–1349. doi: 10.1016/j.cell.2005.09.042. [DOI] [PubMed] [Google Scholar]
  • 15.Menges M., Pavesi G., Morandini P., Bögre L., Murray J.A. Genomic organization and evolutionary conservation of plant D-type cyclins. Plant Physiol. 2007;145:1558–1576. doi: 10.1104/pp.107.104901. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Collins C., Dewitte W., Murray J.A. D-type cyclins control cell division and developmental rate during Arabidopsis seed development. J. Exp. Bot. 2012;63:3571–3586. doi: 10.1093/jxb/ers015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17.Menges M., Samland A.K., Planchais S., Murray J.A. The D-type cyclin CYCD3;1 is limiting for the G1-to-S-phase transition in Arabidopsis. Plant Cell. 2006;18:893–906. doi: 10.1105/tpc.105.039636. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 18.Dewitte W., Scofield S., Alcasabas A.A., Maughan S.C., Menges M., Braun N., Collins C., Nieuwland J., Prinsen E., Sundaresan V., Murray J.A. Arabidopsis CYCD3 D-type cyclins link cell proliferation and endocycles and are rate-limiting for cytokinin responses. Proc. Natl. Acad. Sci. USA. 2007;104:14537–14542. doi: 10.1073/pnas.0704166104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19.Riou-Khamlichi C., Menges M., Healy J.M., Murray J.A. Sugar control of the plant cell cycle: differential regulation of Arabidopsis D-type cyclin gene expression. Mol. Cell. Biol. 2000;20:4513–4521. doi: 10.1128/mcb.20.13.4513-4521.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Sabatini S., Heidstra R., Wildwater M., Scheres B. SCARECROW is involved in positioning the stem cell niche in the Arabidopsis root meristem. Genes Dev. 2003;17:354–358. doi: 10.1101/gad.252503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.ten Hove C.A., Willemsen V., de Vries W.J., van Dijken A., Scheres B., Heidstra R. SCHIZORIZA encodes a nuclear factor regulating asymmetry of stem cell divisions in the Arabidopsis root. Curr. Biol. 2010;20:452–457. doi: 10.1016/j.cub.2010.01.018. [DOI] [PubMed] [Google Scholar]
  • 22.Ortega-Martínez O., Pernas M., Carol R.J., Dolan L. Ethylene modulates stem cell division in the Arabidopsis thaliana root. Science. 2007;317:507–510. doi: 10.1126/science.1143409. [DOI] [PubMed] [Google Scholar]
  • 23.Zhang H., Han W., De Smet I., Talboys P., Loya R., Hassan A., Rong H., Jürgens G., Paul Knox J., Wang M.H. ABA promotes quiescence of the quiescent centre and suppresses stem cell differentiation in the Arabidopsis primary root meristem. Plant J. 2010;64:764–774. doi: 10.1111/j.1365-313X.2010.04367.x. [DOI] [PubMed] [Google Scholar]
  • 24.Zhang W., Swarup R., Bennett M., Schaller G.E., Kieber J.J. Cytokinin induces cell division in the quiescent center of the Arabidopsis root apical meristem. Curr. Biol. 2013;23:1979–1989. doi: 10.1016/j.cub.2013.08.008. [DOI] [PubMed] [Google Scholar]
  • 25.González-García M.P., Vilarrasa-Blasi J., Zhiponova M., Divol F., Mora-García S., Russinova E., Caño-Delgado A.I. Brassinosteroids control meristem size by promoting cell cycle progression in Arabidopsis roots. Development. 2011;138:849–859. doi: 10.1242/dev.057331. [DOI] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Document S1. Supplemental Experimental Procedures, Figures S1–S3, and Table S1
mmc1.pdf (3.2MB, pdf)
Document S2. Article plus Supplemental Information
mmc2.pdf (4.8MB, pdf)

RESOURCES