The hidden half comes into the spotlight: Peeking inside the black box of root developmental phases

Abstract

Efficient use of natural resources (e.g., light, water, and nutrients) can be improved with a tailored developmental program that maximizes the lifetime and fitness of plants. In plant shoots, a developmental phase represents a time window in which the meristem triggers the development of unique morphological and physiological traits, leading to the emergence of leaves, flowers, and fruits. Whereas developmental phases in plant shoots have been shown to enhance food production in crops, this phenomenon has remained poorly investigated in roots. In light of recent advances, we suggest that root development occurs in three main phases: root apical meristem appearance, foraging, and senescence. We provide compelling evidence suggesting that these phases are regulated by at least four developmental pathways: autonomous, non-autonomous, hormonal, and periodic. Root developmental pathways differentially coordinate organ plasticity, promoting morphological alterations, tissue regeneration, and cell death regulation. Furthermore, we suggest how nutritional checkpoints may allow progression through the developmental phases, thus completing the root life cycle. These insights highlight novel and exciting advances in root biology that may help maximize the productivity of crops through more sustainable agriculture and the reduced use of chemical fertilizers.

Keywords: cell fates; developmental transitions; nutritional checkpoints; plasticity; root clock; root development.

Figure 1

Root developmental phase progression. Following meiosis and successful fertilization, seeds originate and protect the embryo. At this stage, stimuli promote seed germination, and patterns of cell division are established whereby the root apical meristem (RAM) emerges to orient cell divisions and growth during the RAM appearance phase. Thus, quiescent-center (QC) cells with the highest identity and the lowest rates of cell differentiation may be observed in this phase. Under adequate timing and environmental conditions, the root-foraging phase begins, with constant increases in cell differentiation and improvements in nutrient uptake. At the foraging phase, nutrient uptake reaches maximum levels (red color at speedometer), whereas minimum levels (green color at speedometer) occur during later developmental stages. Varying with tissue age, species, and root class, and after the maximum foraging time, the entire senescence phase is initiated, during which cell proliferation and differentiation are most likely arrested, and constant patterns of cell death may be observed. Nutrient uptake reaches minimal levels, which in older roots triggers the loss of segments and nutrient remobilization, culminating in root death.

Figure 2

Pathways that promote root developmental phase transitions in Arabidopsis thaliana. (A) The autonomous pathway appears to promote the root-foraging phase, during which the transcription factor MYB36 mRNA is found only in specific cell types, whereas the MYB36 protein induces genes to promote foraging. Similarly, PLETHORA (PLT) expression varies across root zones and is highest around root stem cells, where differential DNA methylation patterns may promote maximum PLT activity. In these cells, RETINOBLASTOMA-RELATED 1 (RBR1) and SCARECROW (SCR) regulate cell fates around QC cells, promoting de novo formative cell divisions. (B) Mobile elements that move among diverse root cell types constitute the non-autonomous pathway. SHORT-ROOT (SHR) moves from the epidermis to the endodermis, where SHR activates SCR to induce the movement of miR165/6, which in turn suppresses the expression of cell-differentiation regulators to release the root-foraging phase. JACKDAW (JKD) activity around QC cells delimits SHR movement in these cells, whereas the expression of genes encoding GLABRA2 (GL2), CAPRICE (CPC), and WEREWOLF (WER) is regulated by JKD, resulting in the mobility of these three genes to give rise to the epidermis. Overall, the non-autonomous pathway sustains the root-foraging phase (from the middle of the phase), vascular tissue formation, and growth. (C) Hormones are essential to organism development, and an emerging hormonal pathway orchestrates root developmental phases. Thus, opposing gradients of auxin and cytokinin regulate patterns of cell proliferation and differentiation. In the root tip, maximum levels of auxin with jasmonates may induce the transcription factor ETHYLENE RESPONSE FACTOR 109 (ERF109) that in turn activates QC cells to promote stem cell fates, particularly around the root tip. Subsequently, a local auxin maximum at the pericycle cells activates CLASS III HOMEODOMAIN-LEUCINE ZIPPER (HD-ZIP III) transcription factors, and increases in auxin and HD-ZIP IIIs lead to the acquisition of xylem identity. As a consequence, ERF109 activity is upregulated at the transition zone according to the balance of jasmonate and auxin, thereby inducing lateral root (LR) formation. Therefore, the hormonal pathway regulates not only the early and later stages of the foraging phase but also entry into the senescence phase. (D) Roots contain cells that vary markedly in ontogeny and require complex genetic circuits to regulate developmental phases, suggesting a periodic pathway. The condensation of AUXIN RESPONSE FACTOR (ARF) transcription factors in the cytoplasm, where they are inactivated, limits auxin responsiveness. Similarly, cyclic cell death of epidermal cells in the transition zone results in periodic LR induction. Meanwhile, the GNOM protein, a vesicle trafficking regulator, is repressed by ARF-GTP ACTIVATING PROTEIN DOMAIN (AGD3), and both proteins regulate the balance between esterification and de-esterification, orienting the turnover of pectin esterification. In the oscillation zone, a root clock is established through a competitive interaction between GNOM and AGD3 that orients the functionality of the LR clock to promote the root-foraging phase. Taken together, evidence suggests that the periodic pathway regulates the overall phases of root development from early until late cell development.

Figure 3

Root developmental phases and plasticity. (A) Roots growing under natural conditions may experience salt stress. Under these conditions, abscisic acid (ABA) triggers suberization of the endodermis, thereby blocking the entry and exit of substances. (B) Roots exhibit a protective structure composed of cutin polyesters (orange caps) that protects the meristem during early development; this structure also protects lateral roots during the foraging phase. Gravitropism has a remarkable effect on EXOCYST70A3 protein activity that in turn modulates the PIN3 transporter, which reorients auxin flux to shift the original elongation plane of the root. Similarly, gravity mediates changes in the amyloplast patterns of root apex cell-group populations, altering the orientation of auxin flow in accordance with PIN2 transporter activity and thereby modifying the root elongation plane. (C) Variations in water potential (Ψw) induce distinct developmental patterns in roots. Higher Ψw promotes optimal activity of AUXIN RESPONSE FACTOR 7 (ARF7), inducing the LATERAL ORGAN BOUNDARIES DOMAIN 16 (LBD16) gene and promoting the root-foraging phase. By contrast, regions of the rhizosphere that are rich in air spaces enhance the SUMOylation of ARF7 (SUMO), thereby recruiting IAA3, repressing LBD16 expression, and arresting root foraging. In these spaces, ABA accumulation impairs cell-identity acquisition, blocking pre-branch site formation. (D) Cell ablation and tissue damage are repaired by means of particular events. Convergent gradients of PLETHORA (PLT) transcription factors and cell regeneration competency are observed over the root regions, and maximum levels of both are found around QC cells. Under optimal growth conditions, PLT may regulate formative cell-proliferation patterns (the common cell cycle with four phases: G₁, synthesis [S], G₂, and mitosis [M]), whereas reactivation of stem cell status requires the re-establishment of the PLT expression gradient (restoration pattern) converging to a gradient of cell regeneration competency at the damaged site. Thus, the restoration pattern shows a divergent cell-division plane for proliferation, in which cell-cycle progression is accelerated by the activation of SHORT-ROOT (SHR), SCARECROW (SCR), and CYCLIN D6;1 (CYCD6;1). This acceleration reflects a shorter duration of the cell-cycle phases (G₁, S, G₂, and M). Consequently, tissue damage (purple circles) activates a hormonal circuit to orchestrate tissue repair. Jasmonates may directly induce ERF115, which regulates stem cell activation and tissue regeneration, and/or induce ETHYLENE RESPONSE FACTOR (ERF) 109 and ERF115, which promote the last steps of tissue regeneration. At the same time, local auxin accumulation at damaged sites promotes the expression of ERF115, which orients stem cell activation and, ultimately, tissue regeneration.

Figure 4

Nutritional checkpoints that govern root developmental phase transitions. Nitrate (NO₃⁻) regulates the activity of the transcription factor TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR1-20 (TCP20), which arrests entry into the root-foraging phase. TCP20 interacts with PLETHORA (PLT) and SCARECROW (SCR) to regulate transcripts such as WUSCHEL-RELATED HOMEOBOX 5 (WOX5), which suggests a role for NO₃⁻ (dotted arrow) in regulating the transition from the embryonic to the root apical meristem (RAM) appearance phase. Low levels of NO₃⁻ drive the preferential gene expression of CLE (CLAVATA3/ESR-related) 1, 3, and 7 peptides in pericycle cells, whereas CLAVATA1 (CLV1) protein is located in the phloem companion cells and represses cell proliferation at lateral roots (LR). LR development is arrested by precise interactions among NO_{3^{− and}} hormones (auxin, cytokinin, and ABA). In general, NO₃⁻ appears to regulate the early root developmental phases, arresting cell differentiation and the root-foraging phase. Phosphate (PO₄³⁻) limitations increase the synthesis/degradation of the transcription factor ROOT HAIR DEFECTIVE 6-LIKE 4 (RSL4), and its protein turnover determines the size of root hairs. Nonetheless, these conditions trigger auxin transport from the root apex to the differentiation zone, where the transcription factors AUXIN RESPONSE FACTOR 19 (ARF19), RSL2, and RSL4 are induced to promote root-branch development and foraging in soil layers. Therefore, phosphorus appears to promote and orient the progression through the foraging phase. In accordance with the general life rule, roots experience aging, culminating in the senescence phase. The molecular mechanisms that program nutritional checkpoints to mediate the root-senescence phase remain poorly documented.