Maintenance Of Stem Cell Niche Research Articles

Histone chaperones play crucial roles in maintenance of stem cell niche during plant root development.

Stem cells in both plant and animal kingdoms reside in a specialized cellular context called the stem cell niche (SCN). SCN integrity is crucial for organism development. Here we show that the H3/H4 histone chaperone CHROMATIN ASSEMBLY FACTOR-1 (CAF-1) and the H2A/H2B histone chaperone NAP1-RELATED PROTEIN1/2 (NRP1/2) play synergistic roles in Arabidopsis root SCN maintenance. Compared with either the m56-1 double mutant deprived of NRP1 and NRP2 or the fas2-4 mutant deprived of CAF-1, the combined m56-1fas2-4 triple mutant displayed a much more severe short-root phenotype. The m56-1fas2-4 mutant root lost the normal organizing center Quiescent Center (QC), and some initial stem cells differentiated precociously. Microarray analysis unraveled the deregulation of 2735 genes within the Arabidopsis genome (representing >8% of all genes) in the m56-1fas2-4 mutant roots. Expression of some SCN key regulatory genes (e.g. WOX5, PLT1, SHR) was not limiting, rather the plant hormone auxin gradient maximum at QC was impaired. The mutant roots showed programmed cell death and high levels of the DNA damage marked histone H2A.X phosphorylation (γ-H2A.X). Knockout of either ATAXIA-TELANGIECTASIA MUTATED (ATM) or ATR, encoding a DNA damage response kinase, rescued in part the cell death and the short-root phenotype of the m56-1fas2-4 mutant. Collectively, our study indicated that NRP1/2 and CAF-1 act cooperatively in regulating proper genome transcription, in sustaining chromatin replication and in maintaining genome integrity, which are crucial for proper SCN function during continuous post-embryonic root development.

The role of CSF1R-dependent macrophages in control of the intestinal stem-cell niche

Colony-stimulating factor 1 (CSF1) controls the growth and differentiation of macrophages.CSF1R signaling has been implicated in the maintenance of the intestinal stem cell niche and differentiation of Paneth cells, but evidence of expression of CSF1R within the crypt is equivocal. Here we show that CSF1R-dependent macrophages influence intestinal epithelial differentiation and homeostasis. In the intestinal lamina propria CSF1R mRNA expression is restricted to macrophages which are intimately associated with the crypt epithelium, and is undetectable in Paneth cells. Macrophage ablation following CSF1R blockade affects Paneth cell differentiation and leads to a reduction of Lgr5+ intestinal stem cells. The disturbances to the crypt caused by macrophage depletion adversely affect the subsequent differentiation of intestinal epithelial cell lineages. Goblet cell density is enhanced, whereas the development of M cells in Peyer’s patches is impeded. We suggest that modification of the phenotype or abundance of macrophages in the gut wall alters the development of the intestinal epithelium and the ability to sample gut antigens.

The Unexplored Role of Intra-articular Adipose Tissue in the Homeostasis and Pathology of Articular Joints.

Intra-articular adipose tissue deposits known as articular fat pads (AFPs) are described to exist within synovial joints. Their assumed role in normal joint biomechanics is increasingly objectivized by means of advanced methods of functional imaging. AFPs possess structural similarity with body subcutaneous white adipose tissue (WAT), however, seems to be regulated by independent metabolic loops. AFP dimension are conserved during extreme WAT states: obesity, metabolic syndrome, lipodystrophy, and cachexia. Hoffa fat pad (HFP) in the knee is increasingly recognized as a major player in pathological joint states such as anterior knee pain and osteoarthritis. HFP contains numerous population of mesenchymal and endothelial progenitors; however, the possible role of mature adipocytes in the maintenance of stem cell niche is unknown. We propose that AFP is an active component of the joint organ with multifunctional roles in the maintenance of joint homeostasis. Endowed with a rich network of sensitive nervous fibbers, AFPs may act as a proprioceptive organ. Adipokines and growth factors released by AFP-resident mature adipocytes could participate in the maintenance of progenitor stem cell niche as well as in local immune regulation. AFP metabolism may be locally controlled, correlated with but independent of WAT homeostasis. The identification of AFP role in normal joint turnover and its possible implication in pathological states could deliver diagnostic and therapeutic targets. Drug and/or cell therapies that restore AFP structure and function could become the next step in the design of disease modifying therapies for disabling joint conditions such as osteoarthritis and inflammatory arthritis.

37.シロイヌナズナの根における幹細胞ニッチの維持に必要な分泌型ペプチドホルモン(口頭発表)

37.シロイヌナズナの根における幹細胞ニッチの維持に必要な分泌型ペプチドホルモン(口頭発表)

PHB3 Maintains Root Stem Cell Niche Identity through ROS-Responsive AP2/ERF Transcription Factors in Arabidopsis

The root stem cell niche, which is composed of four mitotically inactive quiescent center (QC) cells and the surrounding actively divided stem cells in Arabidopsis, is critical for growth and root development. Here, we demonstrate that the Arabidopsis prohibitin protein PHB3 is required for the maintenance of root stem cell niche identity by both inhibiting proliferative processes in the QC and stimulating cell division in the proximal meristem (PM). PHB3 coordinates cell division and differentiation in the root apical meristem by restricting the spatial expression of ethylene response factor (ERF) transcription factors 115, 114, and 109. ERF115, ERF114, and ERF109 mediate ROS signaling, in a PLT-independent manner, to control root stem cell niche maintenance and root growth through phytosulfokine (PSK) peptide hormones in Arabidopsis.

Translesion DNA Synthesis in Cancer: Molecular Mechanisms and Therapeutic Opportunities.

The genomic landscape of cancer is one marred by instability, but the mechanisms that underlie these alterations are multifaceted and remain a topic of intense research. Cellular responses to DNA damage and/or replication stress can affect genome stability in tumors and influence the response of patients to therapy. In addition to direct repair, DNA damage tolerance (DDT) is an element of genomic maintenance programs that contributes to the etiology of several types of cancer. DDT mechanisms primarily act to resolve replication stress, and this can influence the effectiveness of genotoxic drugs. Translesion DNA synthesis (TLS) is an important component of DDT that facilitates direct bypass of DNA adducts and other barriers to replication. The central role of TLS in the bypass of drug-induced DNA lesions, the promotion of tumor heterogeneity, and the involvement of these enzymes in the maintenance of the cancer stem cell niche presents an opportunity to leverage inhibition of TLS as a way of improving existing therapies. In the review that follows, we summarize mechanisms of DDT, misregulation of TLS in cancer, and discuss the potential for targeting these pathways as a means of improving cancer therapies.

In Arabidopsis thaliana Cadmium Impact on the Growth of Primary Root by Altering SCR Expression and Auxin-Cytokinin Cross-Talk.

Cadmium is one of the most widespread pollutant in both terrestrial and marine environment, and its inhibitory effect on plant growth has been largely demonstrated. However, the molecular mechanisms underlying Cd toxicity in plant and mainly in root, as the first organ sensing soil heavy metals, need to be better investigated. To this aim, in the present work we analyzed the growth and the organization of Arabidopsis thaliana primary root in seedlings exposed to Cd (25 and 50 μM) for 8 days starting from germination. Root length, root meristem size, and organization were evaluated together with the behavior of some of the major molecular players in root growth and patterning. In particular, by using different GFP transgenic lines, we monitored: (i) the expression pattern of WOX5 and SCR transcription factors involved in the establishment and maintenance of stem cell niche and in the control of meristem size; (ii) the expression pattern of the IAA-inducible pDR5::GFP reporter, PIN 1, 2, 3, 7 auxin carriers and TCSn::GFP cytokinin-sensitive sensor as relevant components of hormone circuit controlling root growth. We report that Cd exposure inhibits primary root growth via affecting RAM stem cell niche and root radial pattern. At the molecular level, an impairment of auxin maximum accumulation at the root tip, related to a down-regulation and mislocalisation of PIN proteins, and an enhancement of TCSn::GFP cytokinin-sensitive sensor signal is also detected under Cd treatment, thus suggesting an alteration in the homeostasis of auxin/cytokinin signaling. Moreover, and for the first time Cd toxicity on root growth and pattern has been related to a misexpression of SCR transcription factors which is known to interplay with auxin/cytokinin cross-talk in the control of RAM maintenance and activity.

Abstract 1333: Cross talk between mitogen-activated protein kinase signaling cascades and Wnt/β-catenin in HER2+ overexpressing breast cancer cells

Abstract Introduction: The HER2-positive breast cancer is characterized by an amplification of the HER2 gene, resulting in an increase in HER2 presence on the surface of cells and magnification of downstream intracellular signaling. Treatment of HER2+ breast cancer with trastuzumab decreases tumor volume, reduces metastasis and increases overall survival. Unfortunately, HER2-positive breast cancers develop resistance to trastuzumab. The mechanism of resistance may involve the activation of compensatory networks of different cell signaling pathways. The purpose of the study is to examine the crosstalk between HER2/EGFR and Wnt signaling in HER2-positive breast cancer cells. Methods: In order to address this, a HER2-positive breast cancer cell line (SKBR3) was used and induced with EGF. The phosphorylation of p38 and ERK1/2 was determined by Western blot analysis. The cellular localization of proteins of interest was examined using biochemically fractionated lysates followed by western blot analysis. Results: Our data demonstrated an early and transient peak of phospho-p38 at 30min post EGF treatment. Interestingly, both phospho-p38 and phospo-ERK1/2 were detected in the nuclear compartment at this time. β-catenin, the major effector molecule in Wnt signaling, was stabilized 30min post EGF treatment. β-catenin protein levels remained higher in EGF treated cells than untreated cells up to 4h post EGF treatment. It is likely that a stabilized β-catenin following EGF treatment potently modulates a subset of Wnt targets. We are assessing this possibility using chromatin immunoprecipitation based experiments for H3K18ac, H3K27ac, pol2 and β-catenin. Taken together, our data suggests that EGFR signaling promotes Wnt signaling in HER2-positive breast cancer cells. We hypothesize that this promotes cancer stem cell niche maintenance and an EMT phenotype. Citation Format: Miguel Nava, Nwamaka A. Amobi, Yanyuan Wu, Robin Farias-Eisner, Jay Vadgama. Cross talk between mitogen-activated protein kinase signaling cascades and Wnt/β-catenin in HER2+ overexpressing breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1333. doi:10.1158/1538-7445.AM2017-1333

Symplastic communication spatially directs local auxin biosynthesis to maintain root stem cell niche in Arabidopsis

Stem cells serve as the source of new cells for plant development. A group of stem cells form a stem cell niche (SCN) at the root tip and in the center of the SCN are slowly dividing cells called the quiescent center (QC). QC is thought to function as a signaling hub that inhibits differentiation of surrounding stem cells. Although it has been generally assumed that cell-to-cell communication provides positional information for QC and SCN maintenance, the tools for testing this hypothesis have long been lacking. Here we exploit a system that effectively blocks plasmodesmata (PD)-mediated signaling to explore how cell-to-cell communication functions in the SCN. We showed that the symplastic signaling between the QC and adjacent cells directs the formation of local auxin maxima and establishment of AP2-domain transcription factors, PLETHORA gradients. Interestingly we found symplastic signaling is essential for local auxin biosynthesis, which acts together with auxin polar transport to provide the guidance for local auxin enrichment. Therefore, we demonstrate the crucial role of cell-to-cell communication in the SCN maintenance and further uncover a mechanism by which symplastic signaling initiates and reinforces the positional information during stem cell maintenance via auxin regulation.

Indispensable role of Notch ligand-dependent signaling in the proliferation and stem cell niche maintenance of APC-deficient intestinal tumors

Ligand-dependent activation of Notch signaling is required to maintain the stem-cell niche of normal intestinal epithelium. However, the precise role of Notch signaling in the maintenance of the intestinal tumor stem cell niche and the importance of the RBPJ-independent non-canonical pathway in intestinal tumors remains unknown. Here we show that Notch signaling was activated in LGR5+ve cells of APC-deficient mice intestinal tumors. Accordingly, Notch ligands, including Jag1, Dll1, and Dll4, were expressed in these tumors. In vitro studies using tumor-derived organoids confirmed the intrinsic Notch activity-dependent growth of tumor cells. Surprisingly, the targeted deletion of Jag1 but not RBPJ in LGR5+ve tumor-initiating cells resulted in the silencing of Hes1 expression, disruption of the tumor stem cell niche, and dramatic reduction in the proliferation activity of APC-deficient intestinal tumors in vivo. Thus, our results highlight the importance of ligand-dependent non-canonical Notch signaling in the proliferation and maintenance of the tumor stem cell niche in APC-deficient intestinal adenomas.

Reactive Oxygen Species: From Harmful Molecules to Fine-Tuning Regulators of Stem Cell Niche Maintenance.

Reactive Oxygen Species: From Harmful Molecules to Fine-Tuning Regulators of Stem Cell Niche Maintenance.

Association of TGFβ signaling with the maintenance of a quiescent stem cell niche in human oral mucosa.

A dogma in squamous epithelial biology is that proliferation occurs in the basal cell layer. Notable exceptions are squamous epithelia of the human oral cavity, esophagus, ectocervix, and vagina. In these human epithelia, proliferation is rare in the basal cell layer, and the vast majority of cells positive for Ki67 and other proliferation markers are found in para- and suprabasal cell layers. This unique human feature of a generally quiescent basal cell layer overlaid by highly proliferative cells offers the rare opportunity to study the molecular features of undifferentiated, quiescent, putative stem cells in their natural context. Here, we show that the quiescent human oral mucosa basal cell layer expresses putative markers of stemness, while para- and suprabasal cells are characterized by cell cycle genes. We identified a TGFβ signature in this quiescent basal cell layer. In in vitro organotypic cultures, human keratinocytes could be induced to express markers of these quiescent basal cells when TGFβ signaling is activated. The study suggests that the separation of basal cell layer and proliferation in human oral mucosa may function to accommodate high proliferation rates and the protection of a quiescent reserve stem cell pool. Psoriasis, an epidermal inflammatory hyperproliferative disease, exhibits features of a quiescent basal cell layer mimicking normal oral mucosa. Our data indicate that structural changes in the organization of epithelial proliferation could contribute to longevity and carcinogenesis.

A Bistable Switch Mechanism for Stem Cell Domain Nucleation in the Shoot Apical Meristem.

In plants, the stem cells residing in shoot apical meristems (SAM) give rise to above-ground tissues (Aichinger et al., 2012). Hence, the maintenance of stem cell niches is of central importance to a plant's continued growth and development (Fletcher and Meyerowitz, 2000; Gordon et al., 2009). For the flowering plant Arabidopsis thaliana, the genetic determinants of stem cell growth, division, and localization have been identified, and negative feedback between a homeodomain transcription factor, WUSCHEL (WUS), and a receptor kinase, CLAVATA (CLV), is known to play a crucial role in controlling the reservoir of stem cells in the central domain of a SAM. The morphology of plant stems and floral organs is controlled in large part by the size and stability of SAMs, which is controlled, in turn, by spatiotemporal patterns of WUS and CLV expression in meristems. For example, loss of restrictive signals in clv mutants of Arabidopsis leads to enlargement of shoot and floral meristems, resulting in extra floral organs and club-shaped siliques (Jonsson et al., 2005). The size, localization and stability of stem cell domains should be determined, in principle, by the interactions of WUS and CLV proteins, especially by their propensities to diffuse through the domain and by the rates of the molecular reactions that control their activities. Within this paradigm, reaction-diffusion (RD) models of WUS-CLV interactions have been popular mathematical models of SAM dynamics (Jonsson et al., 2005; Hohm et al., 2010; Fujita et al., 2011). In RD models, the spontaneous generation of inhomogeneous distributions of WUS and CLV in SAM domains is usually attributed to mechanisms based on a “Turing” instability (Turing, 1952; Segel and Jackson, 1972). The generic RD equations for spatiotemporal changes in the concentrations, u(x,t) and v(x,t), of two interacting proteins are ∂u∂t=f(u,v)+Du∂2u∂x2, ∂v∂t=g(u,v)+Dv∂2v∂x2, where f(u,v) and g(u,v) are nonlinear functions describing their local chemical interactions. A unique, uniform, steady-state solution, u(x,t) = u0 = constant and v(x,t) = v0, of these equations can become unstable with respect to small, non-uniform perturbations, u(x,t) = u0 + eλt·δu·cos(qx) and v(x,t) = v0 + eλt·δv·cos(qx), δu > Du, generating standing waves of wavelength l ≈ 2π/qcrit in the simulations of the RD system (Gierer and Meinhardt, 1972; Murray, 2003). At present, the diffusive lengths of CLV and WUS in SAMs have not been determined, and there is no evidence to suggest that the Turing condition (diffusivity of CLV >> diffusivity of WUS) is satisfied in the central zone of a SAM.

An in situ hybridization study of Hyaluronan synthase (Has) mRNA in developing mouse molar and incisor tooth germs

Hyaluronan (HA) is a major constituent molecule in most extracellular matrices and is synthesized by Hyaluronan synthase (Has). In the present study, we examined expression patterns of Has1, -2, -3 mRNA in developing mouse molar and incisor tooth germs from embryonic day (E) 11.5 to postnatal day (P) 7, focusing on Hertwig’s epithelial root sheath (HERS) and the apical bud in particular. Has1 mRNA expression was not detected in all tooth germs examined. Has2 mRNA was expressed in the surrounding mesenchyme from E12.0 to 18.0 in both molar and incisor tooth germs, but disappeared after birth. Meanwhile, Has3 mRNA was exclusively expressed within the enamel organ, especially in the inner enamel epithelium (IEE), stellate reticulum (SR), and stratum intermedium (SI) until the early bell stage at E16.0. Has3 mRNA disappeared as IEE differentiated into differentiating ameloblasts (dABs), but remained in SI until the root developmental stage of the molar tooth germ at P7. Has3 mRNA was also expressed in HERS until P7. In incisors, Has3 mRNA was expressed in the apical bud, especially in the transit-amplifying (TA) cell region from E16.0 to P7, and in the papillary layer (PL) adjacent to the mature enamel. These gene expression patterns suggested that Has3 is the main control factor for prenatal and postnatal HA synthesis of the tooth germ, and may in part regulate crown and root formation of the tooth germ, maintenance of stem cell niches in the apical bud as well as mineral transport in PL.

Topoisomerase II-associated protein PAT1H1 is involved in the root stem cell niche maintenance in Arabidopsis thaliana.

PAT1H1, one of the homologues of Topoisomerase II-associated protein, is involved in the maintenance of root stem cell niche through the interaction with NINJA. The root stem cell niche, which possesses four mitotically inactive quiescent cells (QC) and the surrounding mitotically active stem cells, is critical for root development in Arabidopsis thaliana. However, the molecular regulation of the maintenance of root stem cell niche identity is still not fully understood. Here we show that one of the homologues of Topoisomerase II-associated protein, here named as PAT1H1, could regulate root stem cell niche identity. The pat1h1 mutant showed higher frequency of QC cell division and root distal stem cell (DSC) differentiation. With a high expression in roots, PAT1H1 was found to interact with the jasmonic acid (JA) signalling negative regulator Novel Interactor of JAZ (NINJA) and thus regulate root DSC niche identity. Consistent with the active QC cell division, which rarely occurs in wild-type controls, the pat1h1 mutant displayed higher expression of CYCB1 in the root stem cell niche. Together our data reveals that PAT1H1 maintains root stem cell niche stability through the interaction with NINJA and the regulation of cell division.

Cellular and molecular insight into the inhibition of primary root growth of Arabidopsis induced by peptaibols, a class of linear peptide antibiotics mainly produced by Trichoderma spp.

Trichoderma spp. are well known biocontrol agents that produce a variety of antibiotics. Peptaibols are a class of linear peptide antibiotics mainly produced by Trichoderma Alamethicin, the most studied peptaibol, is reported as toxic to plants at certain concentrations, while the mechanisms involved are unclear. We illustrated the toxic mechanisms of peptaibols by studying the growth-inhibitory effect of Trichokonin VI (TK VI), a peptaibol from Trichoderma longibrachiatum SMF2, on Arabidopsis primary roots. TK VI inhibited root growth by suppressing cell division and cell elongation, and disrupting root stem cell niche maintenance. TK VI increased auxin content and disrupted auxin response gradients in root tips. Further, we screened the Arabidopsis TK VI-resistant mutant tkr1. tkr1 harbors a point mutation in GORK, which encodes gated outwardly rectifying K(+)channel proteins. This mutation alleviated TK VI-induced suppression of K(+)efflux in roots, thereby stabilizing the auxin gradient. The tkr1 mutant also resisted the phytotoxicity of alamethicin. Our results indicate that GORK channels play a key role in peptaibol-plant interaction and that there is an inter-relationship between GORK channels and maintenance of auxin homeostasis. The cellular and molecular insight into the peptaibol-induced inhibition of plant root growth advances our understanding of Trichoderma-plant interactions.

CXCR4 and Glioblastoma.

This article focuses on the possible application of antagonists of the G-protein coupled chemokine receptor, CXCR4, for the treatment of glioblastoma and summarises the evidence for CXCR4 antagonism being a viable therapeutic approach. Particular attention is paid to the role of this receptor in cancer stem cell biology, and the maintenance of CXCR4 expression by the glioblastoma key driver mutations. The expression of the CXCR4 receptor, and of its ligand stromal derived factor 1 (SDF-1, CXCL12), is maintained by intracellular pathways via positive feedback loops, and is associated with the epithelial mesenchymal transition (EMT) and the generation and self-renewal of cancer stem cells. SDF-1 and CXCR4 also play a role in the generation and maintenance of the perivascular stem cell niche which contains these cancer stem cells. The available data suggest that most, if not all, glioblastoma cancer stem cells rely on CXCR4 mediated signalling to maintain their phenotype. SDF-1 and CXCR4 are alsoinvolved in many other aspects of brain tumour biology including resistance to radio- and chemotherapy, the migration of cancer cells through the brain, the generation of the tumour blood supply and the recruitment of vascular progenitor cells. These properties suggest that a CXCR4 antagonist would help in the control of this disease.

WOX5 is Shining in the Root Stem Cell Niche.

The WUS-RELATED HOMEOBOX 5 (WOX5) gene is expressed in the quiescent center (QC) to regulate the columella stem cell (CSC) identity. Three recent reports not only show how WOX5 is controlled but also highlight the key role of WOX5 in root stem cell niche maintenance.

A tetratricopeptide repeat domain-containing protein SSR1 located in mitochondria is involved in root development and auxin polar transport in Arabidopsis.

Auxin polar transport mediated by a group of Pin-formed (PIN) transporters plays important roles in plant root development. However, the mechanism underlying the PIN expression and targeting in response to different developmental and environmental stimuli is still not fully understood. Here, we report a previously uncharacterized gene SSR1, which encodes a mitochondrial protein with tetratricopeptide repeat (TPR) domains, and show its function in root development in Arabidopsis thaliana. In ssr1-2, a SSR1 knock-out mutant, the primary root growth was dramatically inhibited due to severely impaired cell proliferation and cell elongation. Significantly lowered level of auxin was found in ssr1-2 roots by auxin measurement and was further supported by reduced expression of DR5-driven reporter gene. As a result, the maintenance of the root stem cell niche is compromised in ssr1-2. It is further revealed that the expression level of several PIN proteins, namely, PIN1, PIN2, PIN3, PIN4 and PIN7, were markedly reduced in ssr1-2 roots. In particular, we showed that the reduced protein level of PIN2 on cell membrane in ssr1-2 is due to impaired retrograde trafficking, possibly resulting from a defect in retromer sorting system, which destines PIN2 for degradation in vacuoles. In conclusion, our results indicated that SSR1 is functioning in root development in Arabidopsis, possibly by affecting PIN protein expression and subcellular targeting.

The impact of Polycomb group (PcG) and Trithorax group (TrxG) epigenetic factors in plant plasticity.

Current advances indicate that epigenetic mechanisms play important roles in the regulatory networks involved in plant developmental responses to environmental conditions. Hence, understanding the role of such components becomes crucial to understanding the mechanisms underlying the plasticity and variability of plant traits, and thus the ecology and evolution of plant development. We now know that important components of phenotypic variation may result from heritable and reversible epigenetic mechanisms without genetic alterations. The epigenetic factors Polycomb group (PcG) and Trithorax group (TrxG) are involved in developmental processes that respond to environmental signals, playing important roles in plant plasticity. In this review, we discuss current knowledge of TrxG and PcG functions in different developmental processes in response to internal and environmental cues and we also integrate the emerging evidence concerning their function in plant plasticity. Many such plastic responses rely on meristematic cell behavior, including stem cell niche maintenance, cellular reprogramming, flowering and dormancy as well as stress memory. This information will help to determine how to integrate the role of epigenetic regulation into models of gene regulatory networks, which have mostly included transcriptional interactions underlying various aspects of plant development and its plastic response to environmental conditions.