A comprehensive understanding of the root growth potential arises from the apical configuration of a primary root – a synonym of taproot root (Baluška et al., 2010; Freschet et al., 2021b). The ability of taproots to penetrate compact soil layers is due to the larger size of the root apex and the rapid elongation behind the root cap (Clowes, 2000), as taproot meristematic cells have a physiological and mechanical advantage over meristematic cells, compared to other root types, e.g., lateral roots (Perilli et al., 2012). Indeed, the pattern of postembryonic root development can be projected through an analysis of the initial cells located in the root apical meristem (RAM) cells (Figure 1; Perilli et al., 2012; Sozzani and Iyer-Pascuzzi, 2014). Ablation of the RAM in water-limited conditions results the formation of a highly branched, shallow root system (Dubrovsky and Gómez-Lomelí, 2003; Shishkova et al., 2013; Drisch and Stahl, 2015), indicating the essential role of a taproot in root system architecture (Dolan et al., 1993; Chapman et al., 2002; Sabatini et al., 2003) for water acquisition from deeper soil layers (Robbins and Dinneny, 2018; Gupta et al., 2020).

There are three unanswered questions remaining in the regard to the taproot root meristems: (1) how does organization and cellular signaling enable a taproot to grow and penetrate deep soil layers, (2) what internal factors enable taproots to grow rapidly and penetrate deep soil layers, and (3) how does soil water limitation induce the vertical growth of taproots. Aside from the unanswered questions above, how much does the genetic control the cell division explain the continued maintenance of root growth and apical dominance of taproot meristems (Perilli et al., 2012; Sozzani and Iyer-Pascuzzi, 2014). Current findings indicate that differences in inter-tissue signaling and the relationship between tissue-types are mostly responsible for matching meristem growth and root topology patterning (Peters and Tomos, 1996). Meristem enlargement, through increased cell division, and the transition of the cells into the expansion zone, occurs not only in response to internal stimuli during plant ontogenic development, but also in direct response to water supply (Benková and Hejatko, 2009; Mira et al., 2017). It seems likely, however, that cell division predominates cell differentiation in taproot meristems over the long-term to prevent the cessation of root growth until roots reach deep soil layers (Shishkova et al., 2008).

Insufficient water availability and associated reduced water uptake by absorptive roots are the main factors contributing to global forest decline (Allen et al., 2015; Choat et al., 2018; Zadworny et al., 2021). Countering drought stress can be achieved by enhancing water acquisition and/or reducing water consumption, while increased root proliferation and taproot elongation increases water uptake from deeper soil layers (Arend et al., 2011; Tuberosa, 2012; Brunner et al., 2015). Mackay et al. (2020) reported that water acquisition in shallow soil layers declines as drought severity increases. Therefore, long taproots can improve water uptake, and help to compensate for increased water usage (Mucha et al., 2018; Skiadaresis et al., 2019), e.g., in oaks that produce a dominant taproot (Osonubi and Davies, 1981; Löf and Welander, 2004; Bréda et al., 2006; Mucha et al., 2018). Deep-rooted plants access water from deep soil layers and transport it to shallow, drier roots, increasing a plants’ ability to survive due to hydraulic redistribution process (Domec et al., 2004; Smart et al., 2005; David et al., 2013). Nevertheless, such watering is rather uncommon as shallow, fine roots are abandoned and die during the dry season in some drought-adapted tree species and grow back when water is available (Montagnoli et al., 2019). This raises the question, whether water limitation contrarily accelerates taproots growth into deeper soils in response to drought. Hormonal induced accumulation of osmoprotectant metabolites enabling root elongation during drought, confirms that this may be the case (Fàbregas et al., 2018). Therefore, a rigorous quantification of the components and molecular mechanisms regulating taproot growth in trees is required, especially among deciduous angiosperms, such as oak and chestnut, as deep taproots may determine resilience to drought. The first step in developing a mechanistic understanding of taproot growth would be to determine the regulatory effect of different phytohormones on cell division in the taproots RAM.

Auxin contributes to the positioning and formation of meristematic cells during organogenesis (Jiang and Feldman, 2010), as well as the retainment of mitotic activity in meristems (Beemster and Baskin, 2000; Galinha et al., 2007; Stepanova et al., 2008), as well as the fast elongation and differentiation of cells (Rahman et al., 2007; Benková and Hejatko, 2009; Ishida et al., 2010). Auxin accumulation in developing RAM cells has revealed that proteins, belonging to the PINFORMED family (PINs; PIN1, 2, 3, 4, 7), are necessary for the formation of an auxin gradient, and regulating the auxin distribution and acropetal transport to the root apex (Blilou et al., 2005). Auxin gradients that induce the expansion of cells and inhibit cell division in the extension zone (Blilou et al., 2005) by the expression of PLETHORA transcription factors (TFs) (Aida et al., 2004), may also regulate taproot elongation. The maintenance of root tip size and growth rate in transgenic Arabidopsis mutants in which PIN genes were silenced, provided evidence that the formation of an internal auxin gradient is indeed correlated with root development (Blilou et al., 2005; Vieten et al., 2005; Dello Ioio et al., 2007), affecting the formation, maintenance, and activity of RAM cells in deciduous trees (Palovaara and Hakman, 2009; Palovaara et al., 2010; Liu et al., 2014; Qi et al., 2020). PINs can significantly impact the rate of root growth and the size of the root tip (Vieten et al., 2005), possibly determining the pattern of taproot elongation in trees. Studies investigating the role of PINs in poplar (Populus), spruce (Picea abies), and pear (Pyrus), have reported a broader and more unique role for these proteins in auxin-controlled root development in trees (Palovaara et al., 2010; Liu et al., 2014; Qi et al., 2020). Some auxin-regulated developmental processes that are unique to woody plants (Liu et al., 2014), may directly affect the root apex expansion (also in taproots) toward wetter areas of the soil (van den Berg et al., 2016). Exploring the auxin regulatory network underlying root development will provide valuable information on the hormonal regulation of the formation and functioning of RAMs and the factors governing meristem size in plants with prominent taproots.

Cytokinins, as well as auxin, are required for the establishment and maintenance of RAM, through the enhanced of mitotic activity of quiescent center cells (QC; Zhang et al., 2013). In contrast to auxin, however, cytokinins control cell differentiation and inhibit root elongation. Studies on cytokinin biosynthesis mutants have shown that cytokinins can regulate the size of RAM. Application of exogenous cytokinins caused a decrease in meristem size, by affecting the rate of meristematic cell differentiation (Dello Ioio et al., 2007). In fact, a reduction in endogenous cytokinin levels in mutants (with a cytokinin level deficiency) results in faster growth of the primary root (Werner et al., 2001). Therefore, repression of cytokinin activity may enhance drought resistance in trees, enabling deeper soil exploitation by taproot elongation (Werner et al., 2001; Calvo-Polanco et al., 2019). Nevertheless, it is essential to determine if cytokinins function alone or interactively with other hormones do contribute to drought tolerance in plants.

Ethylene, generating uneven transverse cell divisions in the QC of a RAM, plays a major role in inhibiting cell proliferation and root growth (Woeste et al., 1999; Schaller and Kieber, 2002; Růžička et al., 2007; Qin et al., 2019). An ethylene dependent pathway involved in inhibiting root elongation was identified in ETHYLENE OVERPRODUCER (eto1) mutants that exhibit enhanced ethylene biosynthesis, relative to wild-type plants, which produce long primary roots (Woeste et al., 1999). Higher root elongation in ethylene resistant ETHYLENE RESISTANT 1 (etr1), ETHYLENE INSENSITIVE2 (ein2), and ETHYLENE INSENSITIVE3 (ein3) mutants also provided evidence that ethylene inhibits root growth (Růžička et al., 2007). The central function of ethylene in relation to root growth allows roots to restrict elongation when needed and extend their growth into deeper soil layers when conditions initiate growth restoration (Negi et al., 2010; Pandey et al., 2021). A lack of alterations in the size of root meristems in these ethylene mutants is consistent with the potential ability of certain taproots to first hold back and then restart growth under specific environmental conditions (Street et al., 2016). The ability to regulate cell elongation through ethylene, cytokinin, and auxin cross-talk may represent an efficient mechanism for directing the position of roots and may also be involved in plant response to drought conditions.

Gibberellins (GA), abscisic acid (ABA), and brassinosteroids (BR) are classes of hormones that can affect root development. Gibberellins act mainly on endodermal cells in root tissues, inducing an expansion of endodermal cells in the root elongation zone, which consequently limits the elongation rate of other root tissues (Ubeda-Tomás et al., 2008). The effect of ABA on root development has been shown to be concentration-dependent: low concentrations of ABA stimulate root elongation while higher concentration deters root formation (Harris, 2015; Rowe et al., 2016; Sun et al., 2020). Low concentrations of ABA enhance the activity of meristematic cells (stem cells) and alter auxin transport and signaling, while the suppressive effect of high concentrations of ABA on root growth are related to its inhibition of cell division in RAMs, as well as cells in the elongation zone (Sun et al., 2020).

Although auxin and ABA affect different aspects of root growth, high levels of ABA reduce auxin levels, which results in root growth inhibition due to the induction of PLT TFs (Yang et al., 2014; Promchuea et al., 2017). Indeed, when the level of drought is too severe, elevated levels of ABA inhibit root growth, which is why ABA is referred to as the stress hormone (Nakashima and Yamaguchi-Shinozaki, 2013). Interestingly, transgenic poplar lines with ectopic expression of abi1 (abscisic acid insensitive1) exhibit an ABA insensitive phenotype, allowing plants exposed to a short-term water shortage an induction of primary root elongation (Sharp et al., 2004). The signaling pathway involving ABA interactions with ethylene, inhibits further primary root growth by increasing ethylene biosynthesis (Sharp et al., 2000; Qin et al., 2019). This suggests that the sensing of low ABA concentrations during episodes of water limitation could promote taproot growth. BR also promote root growth especially during drought periods. The BR biosynthesis maxima in the elongation zone is accomplished by the accumulation of osmoprotectant metabolites, resulting in the elongation of lateral roots and enhancing water uptake (Bao et al., 2004; Fàbregas et al., 2018; Vukašinović et al., 2021). Although examining of a specific hormone has made it possible to understand the mechanism of single hormone biosynthesis, perception, and signaling, the regulation of root development is largely dependent on the interaction of different hormone pathways.

Dynamic root growth is a result of the interaction between hormones affecting biosynthesis, transport, inactivation, perception, signaling pathways and regulating development, maintenance, and RAM function. An increase in auxin levels contributes to lower cytokinins levels. In addition, an increase in the level of cytokinin inhibits the synthesis of auxin (Eklof et al., 1997; Nordstrom et al., 2004; Di Mambro et al., 2017). Cytokinins may also affect, polar auxin transport and the formation of a local auxin gradient during lateral root formation as well as the expression of genes involved in auxin transport (Laplaze et al., 2007; Kuderova et al., 2008). Similarly, root growth is inhibited by the balance between auxin and ethylene. In response to ethylene, auxin accumulates in RAM cells and inhibits cell elongation and cell differentiation, consequently regulating how different components of the root system develop (Casson and Lindsey, 2003). The regulatory role of this balance was demonstrated through the use of mutants in which the biosynthesis, transport, and perception of auxin was affected (Růžička et al., 2007; Stepanova et al., 2007). The inhibition of PLETHORA (PLT) expression by AUXIN RESPONSE FACTOR (ARF), which negatively regulates WUSCHEL RELATED HOMEOBOX 5 (WOX5) transcripts – the driver of stem cell formation – leads to distal stem cell differentiation in RAM (Figure 2; Su et al., 2011). Thus, the molecular interaction between auxin, cytokinins and other hormones controlling meristem development may be applied to the explanation of taproot growth. The question is which combinations regulate root elongation in a similar manner, or if the result varies in taproot vs. lateral root growth. Therefore, to understand the control of taproot growth, there is a need to explore the molecular and genetic mechanisms that regulates root development, through expression and functional analyses.

The establishment of RAM is determined by many factors, including hormone levels, intercellular signaling, and receptors that interact with specific TFs activated in response to internal and external signals (Drisch and Stahl, 2015). Indeed, TFs in plants regulate the transcription of specific genes (Table 1), as well as the responses to external and internal stimuli (Mitsis et al., 2020). For example, MP-dependent TFs regulate auxin transport into cells and play a role in the generation of RAMs, and may control other auxin response genes (Weijers et al., 2006). TFs also play an important role in establishing the QC in embryonic roots and maintaining the QC in mature roots (Forzani et al., 2014). Establishing the QC is accomplished by determining the cell organization required for columella cell identity, and maintaining the undifferentiated status of the QC, which allows the QC to activate root growth to explore new soil spaces, increase root biomass, and enhance water absorption (Motte et al., 2019). Maintaining an area of undifferentiated stem cells in the RAM provides a source of cells needed to produce new roots throughout the lifetime of plants (Sarkar et al., 2007; Drisch and Stahl, 2015).

The ability of taproots to grow deeper may be associated with the maintenance of the columella stem cells in the distal meristem of root tip and regulation of auxin distribution as in lateral roots (Savina et al., 2020). Engaged in the above processes, WOX TFs (WOX 5/7 and WOX11) play a key role in inducing and sustaining primary roots growth, as well as generations of lateral roots, from a primary root (Hu and Xu, 2016; Baesso et al., 2018). For example, in poplar trees, the WOX TFs, WOX 4/5/11 and 12, regulate the development of new lateral roots originating from taproot (Baesso et al., 2020). Tree root systems can extend to considerable widths and depths, thus WOX increasing the ability of a tree to adapt to adverse abiotic and biotic conditions, such as drought or mechanical damage, to which they are exposed continuously. Indeed specific TFs associations have profound effects on plant resistance to drought e.g., the formation of root non-hair cells (Schiefelbein et al., 2014), the differentiation of root epidermal trichoblasts into root hair cells (Clowes, 2000; Ishida et al., 2008), as well as determining the root hair morphology (Bruex et al., 2012). The importance of TFs and the genes they regulate in taproot response under water deficit conditions, however, has not been investigated, and the specific role of TFs in enhancing drought resistance by promoting taproot growth, driven by ABA-regulated auxin transport, remains to be determined (Carlsbecker et al., 2010; Müller et al., 2016).

MicroRNAs (miRNA), along with other growth regulators, form networks controlling gene expressions at a developmental and tissue level, being key for the regulation of root development (Jones-Rhoades et al., 2006; Couzigou and Combier, 2016), also in deciduous and coniferous trees such as Pinus tabuliformis, Larix olgensis, and Poncirus trifoliate (Song et al., 2009; Zhang et al., 2013, 2019; Niu et al., 2015). Particularly, miRNAs play an important role in root morphogenesis, contributing to the regulation of meristem establishment and maintenance, vasculature differentiation, lateral and adventitious root formation, and the regulation of symbiotic interactions (Couzigou and Combier, 2016). The multitude of functional roles played by miRNAs, both in model, annual, and perennial plant species, confirms their integral role in root development (Figure 3). Little is known, however, about the role of miRNAs in the development of taproots in trees. Thus, understanding the role of these RNAs and their interactions with other molecular components, such as genes, TFs, and plant hormones, will assist in the elucidation of the complex pathways that control taproot development and function during foraging for water and nutrients, as overexpression of specific miRNAs increase tolerance to many abiotic stresses by changing root architecture and its adaptive responses to stressful conditions (Zhang, 2015). MicroRNAs and their interactions with other molecular components effectively regulate RAM size and the differentiation of vascular tissue in root, thus, represent a mechanism that could be applied to taproots growth (Khan et al., 2011). A comparison of PHV (PHAVOLUTA) and PHB (PHABULOSA) gene expression in long and short growing roots in miR165/166-resistant mutants indicated that these mutants have a reduced RAM size and a lower level of vascular differentiation than wild-type plants. Hence, miR165/166 regulates root development by controlling RAM size, organ polarity, differentiation of vascular elements, and shape of the root system architecture (Carlsbecker et al., 2010; Couzigou and Combier, 2016).

Hormone signal transduction pathways are also affected by miRNAs. For example, miR390 mediates the miR390–TAS3–AUXIN RESPONSE FACTOR 2/ARF3/ARF4 regulatory pathway, which is involved in auxin signaling, and miR393 represses auxin signaling mediated by its downstream F-box auxin receptor targets, namely, TRANSPORT INHIBITOR RESPONSE 1 (TIR1), as well as AUXIN SIGNALING F-BOX PROTEINS 2 (AFB2) and AFB3 (Yoon et al., 2010; Meng et al., 2011). A negative regulation of ARF TFs by miR160 contributes to the maintenance of adequate auxin homeostasis and further lateral root formation (Wang et al., 2005; Meng et al., 2011), for example. Mutants resistant to miR160, however, exhibited reduced root branching (Couzigou and Combier, 2016). Another miRNA, miR390, expressed in cells located in the region of lateral root initiation downregulates ARF2, ARF3, and ARF4, resulting in the inhibition of lateral root growth (Marin et al., 2010). Furthermore, miR164 acts on the NAC1 TF acts downstream of TIR1 transmitting auxin signals, promotes lateral root emergence and controls lateral root elongation (miR167 acts on ARF7 and ARF19) (Xie et al., 2004; Guo et al., 2005). The modulation of both the primary root and the lateral roots by miRNAs reveals the broad spectrum of action of these growth regulators in root development and function (Gutierrez et al., 2009).

The regulatory function of miRNAs may also affect drought resistance in roots enabling through the expression of drought-responsive genes. In this regard, some miRNAs, such as ABA responsive genes, auxin signaling genes, genes encoding osmolytes, and antioxidant defense-related genes, can promote an accumulation of target mRNAs associated with enhanced stress tolerance (Ding et al., 2013). Notably, many of the miRNAs that respond to drought stress have only been identified in trees such as poplar and larch, and have not been detected in annual plants, such as Arabidopsis or rice. This may indicate a specific role for miRNA in woody plant species with long-term root systems, whether they are broadleaf or coniferous tress. Accordingly, the ability of miRNAs to regulate gene expression in response to drought, may facilitate tree growth and survival under adverse conditions on a long-term basis (Osakabe et al., 2014). The regulation of both, lateral and primary roots growth (Gutierrez et al., 2009), increases the ability to explore of deeper soil layers. Nevertheless, our understanding of the mechanisms and genes controlling taproot growth, development, differentiation, function, and architecture, especially in response to adverse conditions, such as drought, is far from complete.