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Keywords:

  • Morphogenesis;
  • pattern formation;
  • root hair;
  • trichome;
  • transcription factor

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Regulation of Epidermal Cell Differentiation via GL2
  5. Downstream Target Genes of GL2
  6. Perspectives
  7. Acknowledgements
  8. References

Recent plant development studies have identified regulatory pathways for epidermal cell differentiation in Arabidopsis thaliana. Interestingly, some of such pathways contain transcriptional networks with a common structure in which the homeobox gene GLABLA2 (GL2) is downstream of the transactivation complex consisting of MYB, bHLH, and WD40 proteins. Here, we review the role of GL2 as an output device of the conserved network, and update the knowledge of epidermal cell differentiation pathways downstream of GL2. Despite the consistent position of GL2 within the network, its role in epidermal tissues varies; in the root epidermis, GL2 promotes non-hair cell differentiation after cell pattern formation, whereas in the leaf epidermis, it is likely to be involved in both pattern formation and differentiation of trichomes. GL2 expression levels act as quantitative factors for initiation of cell differentiation in the root and leaf epidermis; the quantity of hairless cells in non-root hair cell files is reduced by gl2 mutations in a semi-dominant manner, and entopically additive expression of GL2 and a heterozygous gl2 mutation increase and decrease the number of trichomes, respectively. Although few direct target genes have been identified, evidence from genetic and expression analyses suggests that GL2 directly regulates genes with various hierarchies in epidermal cell differentiation pathways.

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[ Takashi Aoyama (Corresponding author)]


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Regulation of Epidermal Cell Differentiation via GL2
  5. Downstream Target Genes of GL2
  6. Perspectives
  7. Acknowledgements
  8. References

Plant cells differentiate into a wide variety of sizes and shapes both according to their developmental program and in response to environmental stimuli. Because a plant cell is surrounded by its cell wall, the relative position of cells does not change after proliferation, and cell morphogenesis is basically irreversible. Therefore, the morphological differentiation of cells and their disposition patterns are crucial for plants to ensure functional tissue and organ structures. The epidermal cells of Arabidopsis thaliana, including trichomes (hair-like structures on the shoot surface) and root hairs, are excellent subjects for studies on pattern formation and morphological differentiation in plant cells (for recent reviews see Balkunde et  al. 2010; Tominaga-Wada et  al. 2011; Grebe 2012). These studies have revealed not only mechanisms for cell pattern formation, for example, regulation based on cell lineage, positional cues from subepidermal tissues, and lateral inhibition between neighboring cells, but have also elucidated the transcriptional networks that play an integrative role in plant cell differentiation. Interestingly, of such transcriptional networks, those for various epidermal tissues share a common structure consisting of transcription factor genes encoding MYB proteins, bHLH proteins, the WD40 protein TRANSPARENT TESTA GLABRA1 (TTG1) (Walker et  al. 1999), and the homeodomain leucine-zipper protein GLABRA2 (GL2) (Rerie et  al. 1994; Di Cristina et  al. 1996). In the conserved network structure, the GL2 gene is placed furthest downstream, targeted by a trans-activating complex consisting of R2R3-type MYB, bHLH, and TTG1 proteins, and is negatively regulated by R3-type MYB proteins (Pesch and Hulskamp 2009; Schiefelbein et  al. 2009; Balkunde et  al. 2010).

GL2 was originally identified as a gene which, when mutated, resulted in aberrant trichome cell expansion in Arabidopsis, and was also found to be involved in seed coat mucilage production and root hair pattern formation (Rerie et  al. 1994; Di Cristina et  al. 1996; Masucci et  al. 1996). The gene was cloned and shown to encode a homeodomain leucine-zipper transcription factor (Rerie et  al. 1994; Di Cristina et  al. 1996). Its cell-type-specific expression patterns under various genetic backgrounds were determined, revealing its pivotal regulatory role in cell differentiation (Rerie et  al. 1994; Di Cristina et  al. 1996; Masucci et  al. 1996; Hung et  al. 1998; Szymanski et  al. 1998; Lin and Schiefelbein 2001; Costa and Dolan 2003). Although GL2 is assumed to be a bottleneck in the regulatory pathway for cell differentiation in various epidermal tissues, both its physiological outputs in cell differentiation and the downstream target genes of GL2 remain largely unclear. Therefore, the total picture of the regulatory pathway for differentiation into each cell type is also still not clear. In this review, we thus focus on the role of GL2 as an output device of the conserved transcriptional network, and update current knowledge of epidermal cell differentiation pathways downstream of GL2.

Regulation of Epidermal Cell Differentiation via GL2

  1. Top of page
  2. Abstract
  3. Introduction
  4. Regulation of Epidermal Cell Differentiation via GL2
  5. Downstream Target Genes of GL2
  6. Perspectives
  7. Acknowledgements
  8. References

Root epidermis

In Arabidopsis, mature root epidermis is composed of hair (H) and non-hair (N) cell files (Dolan et  al. 1993; Galway et  al. 1994). Each H cell file makes contact with two adjacent underlying cortical cell files (lying over anticlinal cortical cell walls), whereas each N cell file makes contact with a single cortical cell file (lying over periclinal cortical cell walls). Therefore, the nature of Arabidopsis root hair cells is likely determined by a positional cue from the underlayer structure (Balkunde et  al. 2010; Tominaga-Wada et  al. 2011; Grebe 2012). Once the root hair cell morphogenesis has started, cells undergo a series of events, including bulge formation at a position close to the root tip-oriented end in the outer surface, transition to tip growth, and maintenance of tip growth, resulting in a thin cylindrical structure (Dolan 2001; Ryan et  al. 2001; Carol and Dolan 2002). The GL2 gene is expressed preferentially in N cells throughout proliferation and differentiation of root epidermal cells (Masucci et  al. 1996). gl2 mutants form ectopic root hairs in N cell files (Di Cristina et  al. 1996; Masucci et  al. 1996). Based on these facts, GL2 is considered to be a factor in promoting N cell differentiation; in other words, a factor inhibiting H cell differentiation.

Although the complete pathway from positional cue to cell file-specific differentiation is still unclear, genetic and molecular biological studies have revealed the involvement of a transcriptional network in which GL2 acts furthest downstream (Figure  1A). The complex, consisting of the R2R3-type MYB protein WEREWOLF (WER) (Lee and Schiefelbein 1999) or MYB23 (Kang et  al. 2009), the bHLH proteins GLABLA3 (GL3) (Payne et  al. 2000) or ENHANCER OF GLABRA3 (EGL3) (Zhang et  al. 2003), and the WD40 protein TTG1, transcriptionally activates GL2 (Payne et  al. 2000; Bernhardt et  al. 2003; Zhang et  al. 2003; Bernhardt et  al. 2005). Alongside this direct regulation of GL2 by the trans-activating complex, the WRKY transcription factor TRANSPARENT TESTA GLABRA2 (TTG2) (Johnson et  al. 2002) may trans-activate GL2 downstream of the complex, although ttg2 mutations affect neither GL2 expression nor phenotypes in the root epidermis (Johnson et  al. 2002; Ishida et  al. 2007). The complex also trans-activates genes encoding R3-type MYB proteins CAPRICE (CPC) (Wada et  al. 1997) TRIPTYCHON (TRY) (Schellmann et  al. 2002), ENHANCER OF TRY AND CAPRICE1 (ETC1) (Esch et  al. 2004), ENHANCER OF TRY AND CAPRICE2 (ETC2) (Kirik et  al. 2004), and ENHANCER OF TRY AND CAPRICE3/CPC LIKE MYB3 (ETC3/CPL3) (Simon et  al. 2007; Tominaga et  al. 2008). These are expressed preferentially in N cells, and move to H cells where they suppress the trans-activating complex by competing with WER and MYB23, and hence act as negative regulators of GL2 expression (Schellmann et  al. 2002; Bernhardt et  al. 2003; Esch et  al. 2004; Kirik et  al. 2004; Kurata et  al. 2005; Tominaga et  al. 2007, 2008). In contrast, GL3 and EGL3 are expressed preferentially in H cells and move to N cells (Bernhardt et  al. 2005).

image

Figure 1. Regulatory pathways for epidermal cell differentiation.Pathways upstream of GL2 are schematically illustrated for H and N cells in the root epidermis (A), hypocotyl epidermal cells in stoma and no stoma cell files (B), pavement and trichome cells in the leaf epidermis (C), and a cell in the seed coat epidermis (D). Transcription factors acting in each cell are shown. Red arrows and T-bars indicate positive and negative transcriptional regulation, respectively. Blue arrows indicate protein movement between cells. Arrows and T-bars with dashed lines indicate assumed regulation and movement. Cells that strongly express the GL2 gene are shown in green.

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While the gl2 mutant root epidermis loses its cell file specificity for root hair formation, it retains a cell file-specific pattern in cell length and vacuolation timing. Cells are longer and are vacuolated earlier in N cell files than in H cell files, as occurs in cells in the wild-type root epidermis (Masucci et  al. 1996). In contrast, in ttg mutants, cells in the N cell file position obtain all the characteristics of H cells, including short length and delayed vacuolation as well as root hair formation (Galwayet  al. 1994; Masucci et  al. 1996). Thus, it can be concluded that GL2 promotes a part of N cell differentiation (differentiation not to make a root hair) after cell pattern formation, and that the upstream trans-activating complex may regulate all events of N cell development including cell fate determination. Because roots heterozygous for the gl2 mutations gl2–1/+ and gl2–2/+ form ectopic root hairs in N cell files with significantly higher frequencies than do wild-type roots, the gl2 mutations are thought to act in a semi-dominant manner in N cell differentiation (Masucci et  al. 1996). Besides a high-level expression of GL2 in N cells, a low-level expression of GL2 in H cells is thought to be required for normal root hair tip growth because gl2 mutant root hairs branch at a significantly higher frequency than do wild-type root hairs (Ohashi et  al. 2003). This branching phenotype might be related to the gl2 mutant's effect on polysaccharide production in roots (Tominaga-Wada et  al. 2009).

The phytohormones ethylene and auxin are known to promote root hair formation. Roots of the auxin response mutant auxin resistant 2–1 (axr2–1) produce fewer root hairs than do wild-type roots (Wilson et  al. 1990). In contrast, ectopic root hairs develop from N cell files at high frequencies in constitutive triple response 1 (ctr1) mutant roots, in which an ethylene signal is constitutively active (Dolan et  al. 1994). Consistent with this observation, treatments of wild-type Arabidopsis seedlings with auxin (IAA) and the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) induce ectopic root hairs, while treatment with ethylene inhibitors reduce root hairs (Masucci and Schiefelbein 1994; Tanimoto et  al. 1995). Genetic and gene expression analyses have shown that GL2 negatively regulates the actions of ethylene and auxin on root hair formation in its downstream pathways (Masucci and Schiefelbein 1996).

Hypocotyl epidermis

The hypocotyl epidermis shows a cell file-specific pattern in cell differentiation and GL2 promoter activity which is analogous to the pattern in the root (Berger et  al. 1998a; Hung et  al. 1998). Hypocotyl epidermal cell files located outside the anticlinal cell walls of two underlying cortex cell files contain relatively short cells and form stomata (corresponding to H cell files), whereas cell files located outside the periclinal cell walls of a single cortex cell file contain relatively long cells (corresponding to N cell files) (Hung et  al. 1998). In differentiating the hypocotyl epidermis, GL2 promoter activity is stronger in the latter cell files than in the former cell files (Hung et  al. 1998). Similar to root epidermal cell differentiation, file-specific stoma formation in the hypocotyl epidermis is affected by both gl2 and ttg1 mutations, and the difference in cell length is reduced by the ttg1 mutation, but not by the gl2 mutation (Hung et  al. 1998). This indicates that a common transcriptional network containing GL2 is involved in the position-dependent patterning of epidermal cell differentiation throughout the apical–basal axis of Arabidopsis seedlings. In the hypocotyl epidermis, WER and TTG1, and supposedly GL3 and EGL3, are contained in the trans-activating complex, and CPC acts as a negative regulator of the complex. This network structure is very similar to that in the root epidermis. (Costa and Dolan 2003) (Figure  1B). Notably, the position-dependent patterning of GL2 expression begins within protodermal cells at the heart stage of embryogenesis (Berger et  al. 1998b; Lin and Schiefelbein 2001; Costa and Dolan 2003).

Trichomes

The Arabidopsis trichome is a single large cell differentiated from the shoot epidermis through complicated processes, including multiple cycles of endoreduplication, an elaborate series of cell expansions, and cell wall maturation (Hulskamp et  al. 1994). gl2 mutants display aborted trichomes with aberrant cell expansion while they undergo an average of four cycles of endoreduplication like wild-type trichomes (Hulskamp et  al. 1994; Rerie et  al. 1994). The mutant trichomes show no evident outgrowths on the surface of juvenile leaves, and form short spikes or single stalks, instead of the normal three branches, on the surface of adult rosette leaves (Rerie et  al. 1994). In wild-type plants, high levels of GL2 promoter activity have been observed in the entire leaf at early leaf development stages; however, later on, this activity is limited to developing trichomes and cells surrounding early-stage trichomes (Hung et  al. 1998; Szymanski et  al. 1998).

Similar to that in the root and hypocotyl epidermis, the transcription of GL2 is activated by a complex consisting of the R2R3-type MYB protein GLABRA1 (GL1) (Oppenheimer et  al. 1991) or MYB23 (Kirik et  al. 2005), the bHLH proteins GL3 or EGL3, and TTG1 in the trichome cells at all development stages (Figure  1C) (Szymanski et  al. 1998; Schellmann et  al. 2002; Ishida et  al. 2007; Zhao et  al. 2008). R3-type MYB proteins, including TRIPTYCHON (TRY) (Schellmann et  al. 2002), are expressed in the developing trichomes by the trans-activating complex. The proteins move to the surrounding cells and inactivate the trans-activating complex through competition with GL1 and MYB23, leading to the lateral inhibition of trichome formation and the establishment of the trichome formation pattern on the leaf epidermis (Marks and Esch 2003; Esch et  al. 2003, 2004). TTG2 is upregulated by the trans-activating complex, and promotes trichome cell differentiation through an independent pathway of GL2 (Johnson et  al. 2002; Ishida et  al. 2007).

Loss of function of the GL1 or TTG1 genes eliminates trichome development (Hulskamp et  al. 1994; Larkin et  al. 1994), while gl2 mutants form trichomes with aberrant cell expansion and normal nuclear DNA contents (Hulskamp et  al. 1994; Rerie et  al. 1994), indicating that GL2 is a positive regulator for part of the trichome cell differentiation after cell pattern formation, just as it is for N cell differentiation in the root epidermis. This is consistent with the fact that the TTG2 pathway promotes trichome cell differentiation downstream of the trans-activating complex, in parallel with the GL2 pathway. On the other hand, unlike in the root epidermis, in the leaf epidermis GL2 is likely to be involved in cell fate determination and cell pattern formation. This is because transgenic leaves containing the GL2 promoter-driven GL2 transgene, which entopically overexpresses the GL2 function, form more trichomes than wild-type leaves, and frequently develop trichomes adjacent to each other, which rarely occur in wild-type leaves (Ohashi et  al. 2002). Furthermore, gl2–1/+ heterozygous leaves contain fewer trichomes than the wild type and do not display aborted trichomes like those in homozygous mutant leaves (Ohashi et  al. 2002), thus suggesting the involvement of GL2 in trichome cell fate determination.

The phytohormones gibberellin (GA), cytokinin, and jasmonic acid (JA) upregulate trichome formation through an upstream pathway from the trans-activating complex and GL2 (Chien and Sussex 1996; Perazza et  al. 1998; Traw and Bergelson 2003; Greenboim-Wainberg et  al. 2005; Maes and Goossens 2010). GA and cytokinin signaling pathways are integrated by the Arabidopsis transcription factors GLABROUS INFLORESCENCE STEMS (GIS), ZINC FINGER PROTEIN 8 (ZFP8), and GLABROUS INFLORESCENCE STEMS 2 (GIS2), which trans-activate the GL1 gene to promote trichome formation (Gan et  al. 2006, 2007). JA signaling leads to Skp, Cullin, F-box containing (SCF) complex-mediated degradation of jasmonate-ZIM domain (JAZ) proteins, which interact with GL3, EGL3, and GL1 to repress trichome formation (Qi et  al. 2011).

Seed coat epidermis

During seed development, the maternally-derived integuments develop into a seed coat. In Arabidopsis, the outer integument cells differentiate into seed coat epidermal cells, producing a mucilage layer which is visualized by hydration as a halo around the seed (Harris 1991). g12 mutants fail to accumulate the mucilage layer, while other features of the seed coat surface appear unaltered by the mutation (Rerie et  al. 1994). In the seed coat epidermis, the transcription of GL2 is activated by a complex consisting of the R2R3-type MYB protein MYB5 or TRANSPARENT TESTA2 (TT2), the bHLH protein EGL3 or TRANSPARENT TESTA8 (TT8), and TTG1 (Figure  1D) (Zhang et  al. 2003; Western et  al. 2004; Bernhardt et  al. 2005; Gonzalez et  al. 2009; Li et  al. 2009). TTG2 is also positively regulated by the trans-activating complex and is involved in the seed coat mucilage production (Gonzalez et  al. 2009; Western et  al. 2004), implying that GL2 and TTG2 act together in regulating seed coat epidermal cell differentiation.

Downstream Target Genes of GL2

  1. Top of page
  2. Abstract
  3. Introduction
  4. Regulation of Epidermal Cell Differentiation via GL2
  5. Downstream Target Genes of GL2
  6. Perspectives
  7. Acknowledgements
  8. References

Results from intensive expression analyses suggest that a large number of genes are regulated downstream of GL2 during epidermal cell differentiation (Lieckfeldt et  al. 2007; Marks et  al. 2008; Won et  al. 2009; Bruex et  al. 2012). However, compared with its upstream pathways, downstream pathways from GL2 to cell differentiation remain unclear. An Arabidopsis phospholipase D (PLD) gene, PLDζ1, was the first-identified direct target gene of GL2 (Ohashi et  al. 2003). Partial repression of PLDζ1 through inducible RNA interference affects the cell polarity during root hair development, resulting in the random positioning of bulge emergence on the outer surface of H cells and globular root hairs (Ohashi et  al. 2003). The activity of the PLDζ1 promoter is higher in H cell files than in N cell files. This preference disappears in gl2 mutant roots, indicating that PLDζ1 is negatively regulated by GL2 in N cells (Ohashi et  al. 2003). In vitro protein–DNA binding analyses have identified the GL2 binding sequence in the PLDζ1 promoter (Figure  2) (Ohashi et  al. 2003). This sequence includes a sequence similar to the L-1 box which is recognized by ATML1, a homeodomain protein belonging to the same subfamily as GL2 (Abe et  al. 2003).

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Figure 2.     GL2 recognition sequencesGL2-binding sequences in the PLDζ1 and CESA5 promoter regions and two putative GL2 recognition sequences in the XTH17 promoter are shown in the upper portion of the figure. The L-1 box sequence and the relatively conserved sequences around it are marked in red and yellow, respectively. The consensus sequence for GL2 recognition is shown in the lower portion of the figure.

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Although the downstream pathway from PLDζ1 for root hair differentiation is as yet unknown, recent studies on plant signal transduction suggest that it involves a protein kinase cascade containing the 3′-phosphoinositide-dependent kinase PDK1 and AGCVIII kinases (Anthony et  al. 2004; Zhang and McCormick 2009). Unlike animal PDK, which is activated by 3’-phosphoinositides such as PI (3,4,5) P3 (Alessi et  al. 1997), Arabidopsis PDK1 is activated by phosphatidic acid (PA), a product of PLD and PI (4,5) P2 (Anthony et  al. 2004). Depending on PA and not on PI(4,5)P2, PDK1 activates the Arabidopsis AGCVIII kinases including AGC2–1/OXIDATIVE SIGNAL INDUCIBLE1 (OXI1) (Rentel et  al. 2004), which is involved in oxidative stress responses and root hair elongation (Anthony et  al. 2004; Rentel et  al. 2004). These findings suggest that the protein kinase cascade PKD1-AGC2–1/OXI1 mediates the PA signal produced by PLDζ1 to promote root hair elongation. Besides AGC2–1/OXI1, PINOID (PID) (Christensen et  al. 2000; Zegzouti et  al. 2006) and INCOMPLETE ROOT HAIR ELONGATION (IRE) (Oyama et  al. 2002) also belong to the AGCVIII kinase family, and possibly act downstream of PLDζ1. Downstream of GL2, PLDζ1 may accelerate signal transduction, including those signals mediated by reactive oxygen species and auxin which are involved in root hair development.

Polysaccharides in the root cell wall are significantly affected by gl2 mutations, but not by cpc, ttg1, gl1, or gl3 mutations (Tominaga-Wada et  al. 2009). Results from expression analyses of polysaccharide-metabolizing genes strongly suggest that CELLULOSE SYNTHASE5 (CESA5) and XYLOGLUCAN ENDOTRANSGLUCOSYLASE17 (XTH17) are direct targets of GL2 (Tominaga-Wada et  al. 2009). Interestingly, GL2 acts as a negative and a positive regulatory factor of CESA5 and XTH17, respectively. This suggests that GL2 regulates the cell wall structure via the short pathway involving only cell wall-metabolizing genes. The CESA5 and XTH17 promoters contain one and two L1-box sequences, respectively. Yeast one-hybrid analysis confirmed the binding ability of GL2 to the L1-box sequence in the CESA5 promoter (Tominaga-Wada et  al. 2009). The candidate sequences that GL2 recognizes are shown in Figure  2 together with the GL2-binding sequence in the PLDζ1 promoter. They contain several highly-conserved nucleotides outside the L-1 box, suggesting that the whole conserved sequence encompassing the L-1 box is involved in the specific recognition of target genes by GL2.

The ROOT HAIR DEFECTIVE6 (RHD6) gene encodes a bHLH transcription factor belonging to the subfamily VIIIc (Bailey et  al. 2003). Its mutation reduces the number of root hairs, shifts the position of the bulge emergence, and produces a relatively high frequency of epidermal cells with multiple root hairs (Masucci and Schiefelbein 1994). RHD6 is expressed specifically in H cells in the meristem and elongation zones of the wild-type root epidermis. Because the expression spreads to the N cell files in gl2 mutant roots, it has been suggested that RHD6 is suppressed either directly or indirectly by GL2 in N cells (Menand et  al. 2007). However, gl2 rhd6 double mutant roots have more hairs than rhd6 mutants. Therefore, it is likely that RHD6-independent pathways exist downstream of GL2 (Masucci and Schiefelbein 1996). RHD6 is thought to play a role in an auxin- and ethylene-associated process during root hair formation, because the phenotype of rhd6 mutants can be rescued when treated with exogenous auxin or the ethylene precursor ACC (Masucci and Schiefelbein 1994). RHD SIX LIKE1 (RSL1), a gene encoding another bHLH transcription factor belonging to the same subfamily VIIIc, has partially redundant functions with RHD6 in terms of root hair cell differentiation (Menand et  al. 2007). RHD6 and RSL1 may act together downstream of GL2 to positively regulate root hair cell differentiation.

In the seed coat, the rhamnose synthase gene MUCILAGE MODIFIED4 (MUM4), which is required for seed mucilage production (Western et  al. 2004; Oka et  al. 2007), is known to be regulated downstream of GL2. The transcription of MUM4 is suppressed in gl2 mutant seeds during the middle stage of seed development. Interestingly, seed oil levels of gl2 mutants are higher than those of wild-type plants due to suppression of the MUM4 gene (Shen et  al. 2006; Shi et  al. 2012), whereas it is unclear if GL2 is involved in regulating the seed oil level in wild-type seeds.

Perspectives

  1. Top of page
  2. Abstract
  3. Introduction
  4. Regulation of Epidermal Cell Differentiation via GL2
  5. Downstream Target Genes of GL2
  6. Perspectives
  7. Acknowledgements
  8. References

In epidermal cells, the differentiation of which has been found to involve GL2, the complex consisting of R2R3-type MYB, bHLH, and TTG1 trans-activates the GL2 gene. Therefore, the combination of the trans-activating complex and GL2 is considered to be a conserved mechanism for the regulation of epidermal cell differentiation. This idea is applicable to protodermal cells in early embryogenesis shortly after the cellular anatomy of the protoderm and outer ground tissue layer is established (Lin and Schiefelbein 2001). It is also likely that R3-type MYB, as a negative regulator that is mobile between cells, generally helps to establish the epidermal cell differentiation pattern, and that the WRKY transcription factor TTG2 acts downstream of the trans-activating complex in parallel with GL2.

In contrast to the conserved nature of the upstream pathways, no common pathways or regulatory factors have been found downstream of GL2 so far. This may mean that the pathway branches after GL2 to regulate particular genes for the differentiation of each type of cell. In this interpretation, it is noteworthy that GL2 directly regulates the genes encoding the polysaccharide-metabolizing enzymes CESA5 and XTH17 (Tominaga-Wada et  al. 2009), which function in particular cell morphological events. On the other hand, GL2 directly regulates the PLDζ1 gene, whose coding protein produces a phospholipid signal involved in the regulation of various intracellular events including membrane trafficking and intracellular signal transduction (Ohashi et  al. 2003). This indicates that the downstream pathway may be connected to the regulation of intracellular events without additional transcriptional regulation in between. Furthermore, the downstream pathway of GL2 may also contain more transcriptional cascades, because the bHLH transcription factor genes RHD6 and RSL1 have been shown to act downstream of GL2 as positive regulators of root hair development (Menand et  al. 2007). These facts suggest that GL2 directly regulates genes with various hierarchies in pathways for epidermal cell differentiation.

While only a few GL2 direct target genes have been identified so far, genetic and reverse genetic studies have revealed diverse functions of GL2 in epidermal cell differentiation. In the root epidermis, GL2 promotes only a part of N cell differentiation: the escape from root hair formation after N cell fate determination. However, in the leaf epidermis, GL2 is likely to be involved in trichome cell pattern formation and cell fate determination as well as in trichome cell morphogenesis, because entopically additive expression of GL2 was shown to cause adjacent trichomes, which rarely occur in wild-type leaves (Ohashi et  al. 2002). Intriguingly, the expression level of GL2 is thought to be a quantitative constraint on both N cell differentiation and trichome formation, because the number of non-root hair cells is reduced by gl2 mutations in a semi-dominant manner, and because entopically additive expression of GL2 and a heterozygous gl2 mutation increases and decreases the number of trichomes, respectively. GL2 may have a role as an output device that converts an analog signal to an all-or-none response in cell differentiation.

Current evidence suggests a pivotal regulatory role of the GL2 gene in epidermal cell differentiation. Systematic identification of GL2 target genes will help to elucidate the total picture of regulation for epidermal cell differentiation.

(Co-Editor: Chun-Ming Liu)

Acknowledgements

  1. Top of page
  2. Abstract
  3. Introduction
  4. Regulation of Epidermal Cell Differentiation via GL2
  5. Downstream Target Genes of GL2
  6. Perspectives
  7. Acknowledgements
  8. References

We thank Professor Li-Jia Qu and Dr. Tomohiko Tsuge for their help and advice on the manuscript. This work was supported by the Japan-China Joint Research Projects/Seminars between JSPS and the National Natural Science Foundation of China.

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  1. Top of page
  2. Abstract
  3. Introduction
  4. Regulation of Epidermal Cell Differentiation via GL2
  5. Downstream Target Genes of GL2
  6. Perspectives
  7. Acknowledgements
  8. References
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