4 Root cell type GRN adaptations to drought: Impermeabilization
of endodermis and exodermis
The evolution of hydrophobic extracellular biopolymers was a critical
innovation for plant terrestrialization, contributing to permeability
and water transport control. Some of these compounds include complex
phenolic-derived polymers such as lignin, cutin and suberin (Niklas,
Cobb, & Matas, 2017). Lignin generally works in the reinforcement of
secondary cell walls, cutin is commonly found as part of the
impermeabilizing cuticle of aerial parts and was recently reported as
being also part of the root cap cuticle, and suberin is found in many
tissues including specialized root cells, tubers, fruit skin, and seed
coat (Berhin et al., 2019; Niklas et al., 2017; Philippe et al., 2020;
Renault et al., 2017). In general these compounds function in mechanical
support, defence against pathogens and herbivores, and in the control of
the movement of water, nutrient and gases (Niklas et al., 2017; Philippe
et al., 2020; Renault et al., 2017; Pei Wang et al., 2020).
Convergent evolution has also played an important role in the appearance
and shaping of the biosynthetic pathways of some of these biopolymers
across diverging plant taxa. For example, the ancestral green algae and
red-algae have the ability to produce “lignin-like” compounds, leading
to the hypothesis that ancient biosynthesis pathways have been rewired
in the vascular plant lineage (Delwiche et al., 1989; Labeeuw et al.,
2015; Martone et al., 2009). Furthermore, lycophytes and spermatophytes
have independently developed the ability to produce monomers necessary
for lignin biosynthesis and assembly (Renault et al., 2017; Weng et al.,
2010; Weng et al., 2008). It is possible that convergent evolution also
played a role on the appearance of cutin and suberin across distantly
related plant species. Both cutin and suberin share similar initial
biosynthetic steps and lipid precursors, however different enzymes
belonging to the same superfamily and with different mechanisms of
action work on their modification and assembly (Philippe et al., 2020;
Pollard et al., 2008). A recent large-scale comparative genomic study
has revealed that the ability to synthesize precursor molecules of cutin
and suberin emerged prior the evolution of land plants, however the
subsequent steps of the biosynthetic pathway may have evolved
independently across land plants (Cannell et al., 2020). Still, little
is known about how the other parts of the pathway, such as
polymerization and distribution in plant cells, evolved in land plants
(Niklas et al., 2017; Philippe et al., 2020; Pollard et al., 2008).
The diversification of
biosynthetic pathways leading to the production of impermeabilizing
hydrophobic compounds has also contributed to the evolution of distinct
cell types important for adaptation to terrestrial life and for
resilience to drought. For example, the roots of all vascular plants
contain an endodermis surrounding the vascular tissues (Doblas, Geldner,
& Barberon, 2017; Enstone et al., 2002). The endodermis cell layer
forms a diffusion barrier for water, gases and nutrients due to the
presence of two cell wall modifications: the Casparian strip and suberin
lamella (Barberon et al., 2016; Doblas et al., 2017; Seago & Fernando,
2013; Vishwanath, Delude, Domergue, & Rowland, 2015). Casparian strip
is composed of lignin, deposited in the walls of endodermal cells at
their junctions, dividing the layer into outward and inward polarities
and forming an effective barrier to the apoplastic movement of molecules
into the stele and preventing their backflow (Barberon, 2017; Enstone et
al., 2002; D. Roppolo et al., 2014; Daniele Roppolo et al., 2011). The
suberin lamella is a secondary cell wall modification deposited in the
inner surface of the primary cell walls, usually after the Casparian
strip is formed in the mature endodermis (Barberon, 2017; Enstone et
al., 2002). Different from Casparian strip, the suberin lamella may not
form in every root nor in every endodermal cell (the so-called passage
cells) (Andersen et al., 2018; Barberon et al., 2016; Enstone et al.,
2002; Holbein, Shen, & Andersen, 2021). Despite debate in the past
years, the role of suberin lamella as an apoplastic barrier for water
and nutrient uptake from the apoplast to the endodermis cytoplasm has
been demonstrated (Barberon et al., 2016; Ranathunge & Schreiber, 2011;
Peng Wang et al., 2019).
The roots of several species also develop an exodermis below the
epidermis, which is a specialized type of hypodermis with Casparian
bands and suberin lamellae depositions (Enstone et al., 2002; Perumalla
et al., 1990). The exodermis function as a dynamic barrier not only
against water loss under drought and salinity, but also against loss of
oxygen under anoxic conditions, against penetration of ions and heavy
metals, and against pathogen infections (Aloni, Enstone, & Peterson,
1998; Damus, Peterson, Enstone, & Peterson, 1997; Ejiri & Shiono,
2019; Enstone et al., 2002; Líška et al., 2016; Namyslov, Bauriedlová,
Janoušková, Soukup, & Tylová, 2020; Ranathunge, Lin, Steudle, &
Schreiber, 2011; Tylová, Pecková, Blascheová, & Soukup, 2017). At the
same time, the development of exodermis barriers has its downside as it
may impair the uptake of nutrients and interaction with beneficial
microbes (Kamula, Peterson, & Mayfield, 1994). To cope with this
problem, many plant species developed the ability to induce an exodermis
dynamically in response to abiotic stresses, such as drought (Enstone et
al., 2002; Kreszies et al., 2020; Líška et al., 2016; Reinhardt & Rost,
1995; Taleisnik et al., 1999). Interestingly, the development of the
exodermis may vary among closely related species displaying distinct
stress response phenotypes (Ejiri & Shiono, 2019), indicating that this
cell type contributes to plant plasticity and acclimation and may also
help plants to adapt and colonize dry environments.
Regardless of its adaptive role, the evolution of exodermis in plants
still remains untangled. Perumalla et al. (1990) surveyed 181 species
from 53 families of plants from different ecological groups
(hydrophytic, mesophytic, and xerophytic) to determine the presence of
hypodermis with Casparian bands (exodermis). As the majority (156) of
the species assessed presented an exodermis with suberin only
(hypodermis) or with both suberin and lignin, the authors hypothesized
that the presence of a modified hypodermis is ancestral to flowering
plants, and has been retained in many species (Perumalla et al., 1990).
Furthermore, the authors found that festucoid grasses lack Casparian
bands despite presenting cells with similar shape and packing as species
with hypodermal Casparian bands, leading to the hypothesis that their
recent ancestor may have lost the trait. Interestingly in seminal roots
of modern cultivars of barley (a festucoid species) the exodermis fails
to develop even upon severe osmotic stress (Kreszies et al., 2019),
while in wild barley the exodermis is induced in response to osmotic
stress (Kreszies et al., 2020). On the other hand, in other crop grasses
(non-festucoid), such as rice and maize, an exodermis is present and
develops faster in response to stress (Ranathunge, Schreiber, Bi, &
Rothstein, 2016; Schreiber, Franke, Hartmann, Ranathunge, & Steudle,
2005).
Understanding how the exodermis evolved in plants can help in the
identification of the underlying regulatory networks responsible for its
induction in response to drought. To obtain more knowledge about the
evolution of exodermis, we compiled the current information about their
presence in plant species based on literature search (Figure 2) (Bani,
Pérez-De-Luque, Rubiales, & Rispail, 2018; Barrios-Masias, Knipfer, &
McElrone, 2015; Barykina & Kramina, 2006; M. Brundrett, Murase, &
Kendrick, 1990; Calvo-Polanco, Sánchez-Romera, & Aroca, 2014; Damus et
al., 1997; Demchenko, Winzer, Stougaard, Parniske, & Pawlowski, 2004;
Eissenstat & Achor, 1999; Ejiri & Shiono, 2019; Enstone et al., 2002;
Ghanati, Morita, & Yokota, 2005; Kosma, Rice, & Pollard, 2015; Liu et
al., 2019; Perumalla et al., 1990; Ranathunge et al., 2017; Reinhardt &
Rost, 1995; Ron et al., 2013; Schreiber, Franke, & Hartmann, 2005;
Schreiber, Hartmann, Skrabs, & Zeier, 1999; Shiono & Yamada, 2014;
Thomas et al., 2007; Zhang, Yang, & Seago Jr, 2018). Based on this
analysis, the exodermis with suberin first appeared in early land plants
(lycophytes) but it is missing from other seedless vascular plants and
all but one gymnosperm (Damus et al., 1997). Interestingly, four species
in the lycophyte genus Selaginella contain exodermis with lignified
Casparian strips (Damus et al., 1997). Most flowering plants contain an
exodermis with suberization only (hypodermis), while a lignified
exodermis appears in about a third of the species. The scattered
appearances of the exodermal lignification indicates that it has evolved
independently multiple times, suggesting a high evolutionary pressure
and pre-conditioning for the characteristic to arise. Species with no
exodermis have been identified in seven clades (in purple, Figure 2),
and the most parsimonious explanation for presence/absence of exodermis
is the loss of the cell type in these lineages. However, the
evolutionary hypotheses are restricted by the sparse sampling in
families of interest. This is highlighted by the relevant literature
containing contradictions (e.g. pea Bani et al. (2018); Perumalla et al.
(1990); Taleisnik et al. (1999)), likely due to the dynamic nature of
the exodermis.
Evolutionary studies focused on characterizing the exodermis, e.g. by
staining suberin and lignin or using barrier property assays
(Supplementary Table 1) will contribute with important information about
how this cell type has appeared or disappeared multiple times across the
plant lineages. Coupling that with comparative genomics and
transcriptomics of phylogenetically close species (e.g. from the same
family) but with different phenotypes (e.g. non-exodermal, constitutive
and stress-inducible exodermis) will be key to identify the origin of
the regulatory networks and how master regulators underlying exodermis
development and suberization in response to drought evolved. It is
possible that the sporadic appearance of exodermis during plant
evolution was possible through rewiring regulatory networks of Casparian
strips and suberin lamellae formation in endodermis or similar lignin
and suberin biosynthetic pathways from other cell types. The coexistence
of endodermis and exodermis in roots has complicated transcriptomic
approaches, and to investigate the exodermis gene expression and GRNs in
individual species, methods such as laser capture microdissection (LCM)
and translating ribosome affinity purification (TRAP) have been used
(Kajala et al., 2021; Shiono et al., 2014). The exodermis-specific gene
expression patterns and GRNs are a foundation for understanding the
genetic underpinnings of the cell type and enable investigating its
evolutionary paths.
Recent studies are showing the importance of distinct clades of the MYB
transcription factor family as conserved regulators of suberin
deposition in response to osmotic stress in different cell types and in
phylogenetically distant plants, linking their evolution with
colonization of dry terrestrial environments by early land plants
(Capote et al., 2018; Cohen, Fedyuk, Wang, Wu, & Aharoni, 2020; Gou et
al., 2017; Kajala et al., 2021; Kosma et al., 2014; Lashbrooke et al.,
2016; Legay et al., 2016; Shukla et al., 2021; To et al., 2020; Wei et
al., 2020). Two of the possible scenarios are: (1) osmotic
stress-inducible regulation of suberization diversified from
pre-existing developmental pathways, (2) the regulation of suberization
in response to drought was re-activated as plants colonized drier
environments. Taking that into account, further
evolutionary/phylogenetic study of exodermis is needed and selection of
a good clade(s) to dissect the gain/loss events is key for understanding
exodermis development and how it evolves so readily.