The ability to respond to senescence signals and to transition into the reproductive stage develops only in adult plants. Indeed, it has been shown that juvenile
Arabidopsis do not induce senescence symptoms when treated with ethylene [
61]. The length of juvenile growth, however, differs widely between plant species [
29]. This is particularly clear when annual and perennial plants are compared. The juvenile phase in
Arabidopsis, for example, lasts 25 days, whereas woody plants remain in the juvenile phase for years or even decades. Once a plant matures, it enters into reproductive growth and starts to produce flowers and seeds.
2.1. Sequentially Expressed microRNA156 and microRNA172 Decide between Vegetative and Generative Growth
The main regulator responsible for phase transition is microRNA156 [
62]. The level of microRNA156 strongly decreases during the juvenile-to-adult phase transition. The juvenile phase can be prolonged by microRNA156 overexpression, whereas a loss-of-function mutation of microRNA156 forces plant to maturate earlier. In tobacco (
Nicotiana tabacum), microRNA156 overexpression causes the promotion of side shoots and lateral roots development [
63]. These results indicate that microRNA156 is a master regulator of vegetative growth. The second most recognized regulator of plant development is microRNA172. MicroRNA172 is expressed in succession to microRNA156 and increases in mature plants. The homeostasis between these two microRNAs regulates plant maturation and flowering [
62,
64,
65].
Plant life expectancy differs widely between species; nevertheless, the microRNA-dependent maturation mechanism is strongly conserved in flowering species. This conservation has been shown in dicotyledonous plants, such as
Arabidopsis [
65], tomato [
66,
67], tobacco [
68], potato [
68,
69], lotus [
70], cabbage [
71], and alfalfa [
72], and in monocotyledonous maize, rice, and switchgrass [
73,
74,
75,
76]. Additionally, long-living woody species, such as apple tree (
Malus x
domestica) [
77], tea apple (
M. hupehensis) [
78],
Populus x
canadensis,
Acacia confusa,
A. colei,
Eucalyptus globulus,
Hedera helix,
Quercus acutissima [
63], and gymnosperm
Sequoia sempervirens [
79], express microRNA156 to promote vegetative growth in the juvenile phase, while flowering depends on the increase of microRNA172. The sequential expression of these two microRNAs is also visible when juvenile and adult buds or leaves of an individual tree are compared.
MicroRNA156 downregulates SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SBP-like/ SPL)/SQUAMOSA PROMOTER BINDING PROTEIN (SBP) TFs (
Figure 3). In
Arabidopsis, microRNA156 targets the mRNAs of ten out of 16 SPLs [
80]. MicroRNA156 overexpression or loss-of-function mutations of
SPLs phenotypes reveal that SPLs negatively control the initiation rate and number of juvenile leaves, shoot branching, and adventitious root growth while the early stages of flower development are promoted. All these traits are connected to development. Gibberellic acid or floral inductive factors positively stimulate
SPLs expression to levels higher than the microRNA156-set threshold. The microRNA156 level decreases as development progresses and the plant is competent to flower. SPL3 induces the transcription of floral meristem identity genes
LEAFY (LFY),
APETALA1 (AP1), and
FRUITFULL (FUL) by binding to their promoter regions. Overexpressed SPL3, SPL4, and SPL5 are capable of accelerating flowering, while their loss of function does not delay flowering. This suggests that another pathway works in parallel to the microRNA156/SPLs regulatory node [
80].
MicroRNA172 is highly expressed in mature plants and acts through the translational inhibition of
APETALA2 (AP2), an A-class homeotic gene, and AP2-like targets, which include
TARGET OF EAT 1 (TOE1),
TOE2,
TOE3,
SCHLAFMÜTZE (SMZ), and
SCHNARCHZAPFEN (SNZ) (
Figure 3). The overexpression of microRNA172 target proteins represses flowering, while the overexpression of microRNA172 results in an early flowering phenotype [
81,
82].
AGAMOUS (AG), a MCM1-AGAMOUS-DEFICIENS-SRF (MADS)-box TF, is a C-class homeotic gene involved in floral patterning. Its activity is involved in the determination of stamens in the third whorl of a flower and carpels in the fourth whorl of a flower [
83]. A closer investigation into young floral primordia reveled that
AG expression, limited to the center of the developing flower, overlaps with microRNA172 expression. MicroRNA172 post-transcriptionally limits
AP2 expression to the outer whorls of developing flower, where AP2 determines sepals and petals development and restricts AG function. Therefore, microRNA172 plays a dominant role in
Arabidopsis flower patterning [
84]. AP2 binds to the second intron of
AG to act in its transcription repression [
85,
86]. Similarly, TOE3 binds to the second intron of
AG to repress its expression. Moreover,
TOE3 is activated by SPL3, binding to its promoter, and in this way,
TOE3 overcomes the downregulation driven by microRNA172. Consequently, its transcript level gradually increases during development. Additionally, the SPL3–TOE3 interaction links regulatory nets of microRNA172 and microRNA156. The role of TOE3 in flower patterning is not yet established [
82]. Jibran et al. (2017) proposed that AG functions not only in flower initiation, but also in flower development and senescence [
87]. Binding AG to the
ANTHER DEHISCENCE 1 (DAD1) promoter induces
DAD1 gene expression to provide substrates for jasmonic acid (JA) biosynthesis. In a senescing flower, JA regulates stamen dehiscence, sepal yellowing, and perianth abscission. Such a phenomenon is postulated to be responsible for the ephemeral phenotype of
Arabidopsis flowers [
87]. Ephemeral flower senescence is independent of pollination and concurrent ethylene synthesis; thus, the flower lasts less than one day [
88].
Juvenile-to-mature phase transition can be reversed during grafting. The function of microRNA156/SPL3 and microRNA172/AP2 in the process is crucial. While adult shoots of
S. sempervirens rejuvenate when grafted onto juvenile rootstocks, the sequential expression of microRNA156 and microRNA172 is reversed [
79]. Rejuvenated shoots have similar levels of the two microRNAs as juvenile shoots, as well as similar physiological characteristics such as rooting, photorespiration rates, or abscisic acid (ABA) and ethylene levels.
The developmental transition of
Physcomitrella patens is reflected in the switching from the two-dimensional growth of a creeping protonema to the upward growth of a leafy gametophore [
89]. The regulatory network of the developmental transition is only partially conserved in this gametophyte-dominated plant. MicroRNA156 downregulates SPL/SBP mRNAs, while the microRNA172/AP2 pathway is absent. Moreover, microRNA156 stimulates leafy gametophore development, promoting phase transition, which is in opposition to its function in the prolongation of the juvenile stage in angiosperms. The highest level of microRNA156 is detected during bud formation by a protonema, an intermediate stage between a protonema and a leafy gametophores. Further in development, microRNA156 is downregulated, whereas the levels of its
P. patens targets SBP3, SBP6, and SBP13 increase [
89].
The discovery that the decrease in the microRNA156 level ends the vegetative growth stage in angiosperms was a break-through in understanding the juvenile-to-adult transition. The source of the signal and mechanism initiating the time-based decrease of microRNA156 is not well known. In
Arabidopsis, maize, and
Nicotiana benthamiana, the signal probably derives from leaf primordia and inhibits microRNA156 expression [
90].
Arabidopsis roots and cotyledons remain neutral during such signaling. In
Arabidopsis, microRNA156 expression is repressed by microRNA159 overexpression, and vegetative growth is shortened [
91]. Accordingly, the loss of microRNA159 results in an increase in microRNA156 and prolonged juvenile growth. MicroRNA159 decreases
MYB33 expression, the transcription factor that binds to the promoter regions of microRNA156 genes
MIR156A and
MIR156C, as well as to
SPL9. The binding of MYB33 to the gene promoter regions of genes
MIR156A and
MIR156C and to SPL9 facilitates their expression (
Figure 3). Therefore, the role of MYB33 in the maintenance of the balance between microRNA159 and microRNA156 levels is not clear [
91]. MicroRNA156 and microRNA172 expression is regulated by MADS TFs. SHORT VEGETATIVE PHASE (SVP) MADS TF binds to
MIR172’s promoter, lowering its expression [
92,
93]. Moreover, AGAMOUS-like 15 (AGL15) and AGL18 interact together to facilitate the expression of
MIR156 [
94]. Altogether, the binding of SVP to MIR172 or the AGL15/AGL18 complex to
MIR156 promoters leads to flowering retardation. MicroRNA156 and microRNA172’s sequential expression is regulated by POLYCOMB REPRESSIVE COMPLEX1 (PRC1) components:
A. thaliana B lymphoma Moloney murine leukemia virus insertion region1 homolog (AtBMI1-PRC1) and EMBRYONIC FLOWER (EMF1-PRC1) [
95].
MIR172 and
SPLs are inhibited by EMF1-PRC1, which allows plants to stay in the juvenile phase. While a plant matures,
MIR156 is repressed by AtBMI1-PRC1. PRC1 components downregulate gene expression through the introduction of histone-modifying marks. Indeed, the deposition of H3K4me3 at
MIR156A and
MIR156C is correlated with a temporal change in the expression of microRNA156 [
96,
97].
2.2. MicroRNA160, microRNA167, and microRNA390 Guard Auxin Signaling
Auxin suppresses plant senescence by acting in many developmental pathways. In
Arabidopsis, microRNA160 targets ARF10, ARF16, and ARF17 mRNAs from the 23
AUXIN RESPONSE TF genes [
57], whereas microRNA167 is complementary to ARF6 and ARF8 mRNAs (
Figure 3) [
57,
58]. MicroRNA160 levels vary in
Arabidopsis leaves. First leaves highly express microRNA160, while in leaves that emerge later and are therefore younger, the microRNA160 expression is lower [
98]. The disruption of microRNA160-driven regulation of ARF17 in
Arabidopsis causes a number of severe developmental defects in leaf, root, and flower morphology, with the most prominent effects including reduced plant size, accelerated flowering, reduced fertility, and sometimes premature death (before flowering) [
99]. MicroRNA160-ARF17 regulation is important for proper auxin signal distribution in aerial parts of the plant. In rice,
OsARF18 is expressed mainly in leaves and in spikes [
100]. The deregulation of
os-microRNA160a/b-OsARF18 modules by mutation in the microRNA binding site causes pleiotropic developmental defects. This phenotype includes dwarfing, a lower number of tillers, shorter and rolled leaves, and flower and seed abnormalities. Auxin induces the expression of
OsMIR160a and
OsMIR160b, as well as
OsARF18, whereas OsARF18 represses
os-microRNA160a/b expression in a sort of reversed feedback-loop. The importance of microRNA 160 is broader than the regulation of ARFs, because simple overexpression of ARFs has no phenotype [
99].
ARF2, ARF3/ETT, and ARF4 are targeted by ta-siRNAs derived from
TAS3 non-coding RNA (TAS3-ta-siRNAs) (
Figure 3) [
59]. MicroRNA390 sets TAS3 phasing in TAS3-ta-siRNAs biogenesis [
59,
101,
102]. The microRNA390-TAS3-ta-siRNAs-ARF2/3/4 regulatory module is highly conserved in land plants except for lycophytes. Normally, TAS3-ta-siRNAs delay juvenile-to-adult transition, as the neutralization of the TAS3-ta-siRNAs binding site in ARF3 or ARF4 mRNA accelerates phase change in
Arabidopsis [
103,
104]. Similarly, microRNA390 overexpression delays leafy gametophore formation in
P. patens [
89]. The
Arabidopsis phase-change phenotype is ARF3/4 dosage-dependent [
103]. Nevertheless, the TAS3-ta-siRNAs, ARF3, and ARF4 RNA levels do not change during
Arabidopsis development. Therefore, microRNA390-TAS3-ta-siRNAs-ARF3/4 probably secures a threshold below which plants do not enter the mature life stage [
104]. Moreover, ARF2 is recognized as positive regulator of leaf senescence and a major player in auxin-dependent leaf longevity. Lim et al. (2010) identified an
ore14/arf2 Arabidopsis mutant with increased sensitivity to auxin [
105]. This is caused by impaired repression of auxin signaling mediated by ARF2. The mutation caused a delay in all the senescence traits tested, such as chlorophyll content, photosystem II photochemical efficiency, membrane ion leakage, and the expression of senescence-associated genes.
Flower senescence largely depends on microRNA390-TAS3-ta-siRNA-regulated ARF2.
ARF2 mRNA and proteins are accumulated during senescence. Consequently, an
arf2 T-DNA insertion mutation is characterized by the delayed yellowing of rosette leaves and the longer living plant sets flowers later and in higher number. More interestingly, the depletion of ARF2 results in delayed abscission of floral organs, and sepals remain green and turgid when abscised. Additionally, siliques dehiscence emerges later [
106].
2.3. Temporally Acting microRNA396 and microRNA164 Regulate Leaf Longevity
An individual plant produces leaves of various shapes. Leaf morphology depends on plant age and differs between juvenile and adult plants or individual branches in the case of some woody species. In
Arabidopsis, juvenile leaves are characterized by round, flat, and smooth blades with long petioles, whereas mature leaves have elliptical, hyponastic, and serrated blades with shorter petioles [
107]. The pattern of leaf abscission is another morphological trait that differentiates juvenile and adult shoots of trees. Juvenile branches retain their leaves until spring, while adult branches drop their foliage in the fall.
Leaf growth begins on the sides of the shoot apical meristem [
108,
109,
110]. A rod-shaped leaf primordium grows to generate flat lamina. Initially, cells proliferate throughout the primordium. Then, the cell divisions are limited to the leaf base, and the proliferation arrests. Afterwards, leaf growth is limited to the expansion of the cells [
111]. Eventually, the mature leaf becomes a source of assimilates to the whole plant. Finally, the last stage of leaf development is senescence.
MicroRNA396 limits growth-regulating factors (GRF) expression (
Figure 3) [
112]. It targets seven out of nine
Arabidopsis GRFs. The microRNA396 level positively correlates with the age of the leaves; it increases while leaf cells proliferate and after the proliferation arrest [
113,
114,
115]. As a consequence, microRNA396 limits GRFs expression to the basal part of a young leaf blade containing proliferating cells, and later, it arrests GRFs activity throughout the leaf. Hence, the regulation of GRFs by microRNA396 influences the duration of leaf cells proliferation and, consequently, leaf size. Another result of microRNA396’s regulation of GRFs is the control of leaf longevity [
115].
Arabidopsis plants expressing the modified
GRF3 gene, which is insensitive to microRNA396-driven downregulation, show not only increased leaf size, but also a late-senescing phenotype. The delayed senescence does not depend on the prolonged duration of the leaf cell proliferative phase, because the late-senescing phenotype is not present when microRNA396-insentisitve GRF3 is expressed only during the early stages of leaf development. Hence, late in development, GRFs expression influences leaf longevity but not the organ size [
115].
Another temporally regulated microRNA is microRNA164. Its expression gradually decreases in
Arabidopsis leaves during aging [
116]. MicroRNA164 negatively regulates
ORESARA1 (ORE1/AtNAC2/ANAC092) TF expression (
Figure 3). ORE1 mRNA downregulation takes place in younger leaves and is released during aging due to the decline of microRNA164 [
116,
117]. Moreover, the increase of
ORE1 expression is directly driven by the binding of ETHYLENE INSENSITIVE3 (EIN3) to its promoter region [
118,
119]. EIN3 is a key TF stabilized by ETHYLENE INSENSITIVE2 (EIN2/ORE2/ORE3).
MIR164 transcription is inhibited by the direct binding of EIN3 to its promoter. The EIN2-EIN3-ORE1/MIR164-ORE1 pathway is the effector of a signaling cascade of ethylene, a hormone known to accelerate senescence. ORE1 is a positive regulator of senescence and age-related cell death in
Arabidopsis leaves.
ore1 mutants’ late-senescence phenotype is characterized by delayed cell death and, therefore, the loss of chlorophyll content, loss of photochemical efficiency (Fv/Fm), increase in membrane ion leakage, and increased cysteine protease-encoding
senescence-associated gene 12 (SAG12) expression [
120]. Moreover,
ein3 mutants are characterized by a stay-green phenotype. Conversely, cell death occurs earlier during leaf development in
microRNA164a/b/c mutants. The
Arabidopsis ethylene-induced senescence signaling pathway includes several members of NAM/ATAF1,2/CUC2 (NAC) TFs [
118,
119]. Except
ORE1, EIN3 induces
NAC-LIKE ACTIVATED BY APETALA3/PISTILLATA (AtNAP/ANAC029) expression, whereas EIN2 promotes the expression of
ANAC019,
ANAC047,
ANAC055, and
ORS1/ANAC059. EIN3-induced ORE1 and AtNAP activities have partially additive functions in age-dependent and artificially induced leaf senescence. ORE1 induces
ANAC087 and
ANAC102 expression, while ORE1 and AtNAP activate three common NAC TF genes
ANAC041,
ANAC079, and
VND-INTERACTING2 (VNI2). However, VNI2 is known as a negative regulator of leaf senescence [
121]. The wide cascade of ethylene-induced NAC TFs assures elasticity in promoting senescence. Notably, microRNA164-regulated ORE1 is directly involved in leaf de-greening during senescence. ORE1 binds to the promoter regions of
STAY-GREEN1 (SGR1/NYE1), chlorophyll
b reductase (
NYC1/NOL), and pheophorbide a oxygenase (
PaO)—proteins essential in chlorophyll catabolism—to facilitate their expression [
120]. In the case of NYE1 and NYC1, ORE1 function is additive to EIN3, which also binds to the promoters of the genes encoding the same proteins but in a separate promoter site. Moreover, ORE1 stimulates ethylene biosynthesis in a feed-forward manner by inducing 1-aminocyclopropane-1-carboxylicacid synthase (ACS2).
2.4. Cell Divisions and JA Biosynthesis Are Linked to Leaf Senescence by microRNA319 Regulatory Modules
MicroRNA319 regulates five TEOSINTE BRANCHED/CYCLOIDEA/PCF TFs (TCPs) known as negative regulators of cell differentiation and positive regulators of senescence (
Figure 3) [
3]. The function of miRNA319 (miR-JAW) as a negative regulator of plant senescence was first shown in a microRNA319a-overexpressing
Arabidopsis mutant (
jaw-D). This mutant shows the delayed leaf senescence/prolonged juvenile-phase phenotype that is reversed by the addition of JA [
122]. Mild overexpression of microRNA319-non-targeted TCP4 in
jaw-D also relieves the phenotype [
123]. Schommer et al. (2008) linked the microRNA319-TCP regulatory module to the JA biosynthesis pathway [
122]. TCP4 induces
LIPOXYGENASE2 (LOX2) expression, encoding a chloroplast-localized enzyme dedicated to the JA biosynthesis pathway [
122,
124,
125]. The TCP4 binding motif in the promoter region of
LOX2 links this pathway to development, as TCP4 does not react to environmental stimuli. A consequence of this is a higher JA biosynthesis rate in response to developmental changes. JA is a positive regulator of senescence. Nevertheless, JA on its own is not essential for senescence to occur, as JA biosynthesis (
aos, opr3, LOX2-RNAi) or JA-signal transduction defective mutants (
coronatine insensitive1, coi1) are not impaired in senescence. Hence, the endogenous JA level is not a senescence-limiting factor. Exogenously applied JA accelerates senescence in
jawD, as well as wild type (WT) plants, while JA-signal transduction defective mutant
coi1 stays insensitive to JA. It cannot be ruled out, however, that JA promotes senescence as an exogenous signal and that, therefore, it is a way of communicating between plants to coordinate developmental processes. The role of type II TCPs in cell proliferation arrest is better understood. TCP4 promotes the expression of
CYCLIN-DEPENDENT KINASE INHIBITOR 1 (ICK1)/KIP RELATED PROTEIN1 (KRP1) working in a pathway inhibiting the progression of the cell cycle [
126]. MicroRNA319-regulated TCP4 directly induces the expression of
MIR396b in
Arabidopsis. The binding of TCP4 to the
MIR396 promoter results in differentiated expressions of microRNA396 along the leaf with the highest level at the distal parts of the organ. The accumulation of this microRNA increases with leaf age. The role of the expressed microRNA396b is to withhold cell proliferation by targeting GRF TFs [
126,
127].
2.7. MicroRNAs in Fruit Ripening and Senescence: Agronomical Traits
Senescence, as a process, according to the source‒sink hypothesis, redirects nutrients and assimilates from leaves, which have fulfilled their role as a source, to developing fruits. The involvement of microRNAs in the mobilization of resources is made obvious by the different and often opposite expression trends of many microRNAs when senescing
Arabidopsis leaves and siliques have been compared [
117]. Therefore, the manipulation of leaf senescence can be used as a tool to improve the yield, quality, or shelf life of fruits and grains. Tomato,
Solanum lycopersicum, is a fleshy, climacteric fruit model plant, as well as an economically important crop plant. Indeed, knocking down the expression of
SlORE1S02, an
AtORE1 ortholog with a disrupted microRNA164 hybridization site, led to delayed senescence, which was evidenced by a stay-green phenotype [
132]. Moreover, as a result, the fruit yield was higher and had an increased soluble sugar content.
Tomato fruit coloration largely depends on microRNA156 expression. The level of microRNA156 decreases during tomato ripening, and the overexpression of microRNA156 results in the pale red coloration of ripened fruit [
133,
134]. A natural epigenetic mutation in
COLORLESS NON-RIPENING (CNR) of tomato, a SBP-box TF gene whose mRNA is targeted by microRNA156, has been identified.
CNR is upregulated in fruit during the breaker stage, consequently, red coloration in the fruits of the CNR mutants is absent. Other important targets for tomato ripening and softening include endo-1, 4-beta-glucanase, pectate lyase, and beta-galactosidase, which are targeted by microRNA396, microRNA482, and novel microRNAZ7, respectively [
134]. Importantly, these three microRNAs are upregulated during the breaker stage of tomato. In contrast, ethylene treatment, which is responsible for the red coloration of climacteric fruits, decreases the expression of these microRNAs and microRNA156. Microtranscriptome analysis reveals a global decrease of known conserved and known non-conserved microRNAs during tomato fruit ripening from the mature green to the red stage [
134]. In
Solanaceae, microRNA1917 is involved in pedicel abscission [
135]. MicroRNA1917 targets tomato
CONSTITUTIVE TRIPLE RESPONSE1-LIKE4 (CTR4) splicing variant 3 (SlCTR4sv3), which is specifically expressed in the abscission zone.
SlCTR4 regulates ethylene signaling. Sly-miR1917 overexpression changes tissue-specific ethylene responses due to increased ethylene synthesis. In adult plants, this results in accelerated pedicel abscission and fruit maturation. In other plants, CTR expression is induced in ripening fruits and in cut flowers during storage, but the microRNA1917-regulatory module is not conserved out of
Solanaceae.
MicroRNAs known to regulate
Arabidopsis development and senescence were identified in non-climacteric fruits during ripening or postharvest senescence. Strawberry
Fragaria ananassa is a model plant for non-climacteric fruits. The edible part of a strawberry is the receptacle. Initially, it grows under the hormonal control of achenes [
136]. Later, the color change is driven by the action of several TFs. The source-to-sink transition during the color change of the receptacle is driven by the gibberellin- and abscisic acid-regulated MYB family TF (GAMYB). GAMYB binds to the promoter regions of GA-responsive genes to activate their expression. In strawberry,
FaGAMYB knockdown results in retained photosynthetic activity accompanied by a decrease in anthocyanin and sucrose concentration [
137,
138,
139]. The role of
FaGAMYB in the red coloration of the strawberry receptacle can be linked to
FaMYB10 and
FaMYB1 expression induction, as MYB TFs promote anthocyanin biosynthesis. Generally, the downregulation of FaGAMYB results in the changed expression of 2624 genes; therefore, its role can be considered to be dominant during the ripening of the strawberry receptacle. The role of microRNA159 in the process has been well described [
137,
140]. A high concentration of gibberellic acid downregulates
fan-miR159a, one of the two microRNA159 that target GAMYB. Low levels of the two microRNA159 overlap with the peak of GAMYB expression. In strawberry, high-throughput sequencing of small RNAs revealed several microRNAs and their targets, which act during the postharvest senescence of the receptacles [
141]. These microRNAs are microRNA156, microRNA160, microRNA164, microRNA167, microRNA172, and microRNA390. A comparative analysis of microRNAs and their targets in fruits was also conducted for the spontaneous, late-ripening mutant of sweet orange
Citrus sinensis Fengwan and wild-type Fengjie 72-1 [
142]. From the whole pool of conserved microRNAs, only 15% were differentially expressed between the two cultivars. As expected, among them were microRNA156 and microRNA159. Therefore, the SPLs and GAMYBs pathways are differentially regulated between these two cultivars. Interestingly, 38% of the novel citrus microRNAs were deregulated between the differently ripening cultivars. This suggests that species-specific microRNAs are responsible for observed cultivar variations, which opens new pathways for agronomically important research. Unusual targets of conserved microRNAs with important roles in fruit storage have also been identified. The pink-to-brown change of the peel coloration of
Litchi fruit is a commercially recognized, senescence-associated trait [
143]. Important for
Litchi fruit storage, the positive regulators of the anthocyanin biosynthesis pathway are MYB TFs [
138,
139]. In
Litchi, one of the MYB TFs is targeted by microRNA858. The expression of microRNA858 increases during postharvest senescence in
Litchi, which may negatively influence the pink coloration of the peel. The flavonoid biosynthesis enzyme chalcone isomerase is regulated by microRNA396 in
Litchi. MicroRNA396 is upregulated during room temperature storage, while cold room temperatures decrease its expression. Another interesting target of
Litchi microRNA396 is the mRNA of the proteolytic enzyme CYSTEINE PROTEASE1 CP1. Proteolysis leads to cell death and therefore is an important part of the late senescence of
Litchi fruit. The CP1 mRNA level fits to the expression pattern of its regulatory microRNA. These targets have not been previously reported; nevertheless, such additional regulatory nodes can be a part of species-specific posttranscriptional regulation [
143].
2.8. MicroRNA156 and microRNA172, the Main Drivers of Plant Development, Also Act in Global Proliferative Arrest
Grains, legumes, and other economically important plants, as well as
Arabidopsis, are characterized by a single reproductive cycle, semelparity, after which the plant dies. This is due to synchronized arrest of the meristem cell divisions known as global proliferative arrest (GPA). GPA is followed by grain filling and occurs after a given number of fruits is set. Therefore, GPA is assumed to target the allocation of resources to growing seeds. Age-control of shoot apical meristem (SAM) divisions by GPA is also regulated by microRNAs (
Figure 3). Balanza et al. (2018) identified
ful-1 (Ler) and
ful-2 (Col) mutants with prolonged activity of SAM and, consequently, greatly delayed GPA [
144]. Interestingly, the elimination of the microRNA172 binding site in AP2 enhances the phenotype of the
FUL defective mutant in that the GPA is prevented and the meristem stays active. The expression of AP2 that cannot be regulated by microRNA172 in senescent wild-type plants that have undergone GPA restores SAM activity, which makes new flowers and fruits.
FUL and microRNA172 are overexpressed during inflorescence development to inhibit
AP2 and
AP2-like expression. At the same time, the microRNA156 level drops, and upregulated SPL factors strongly induce
FUL [
144]. AP2 is a positive regulator of the
WUSCHEL (WUS) gene, a homeodomain transcription factor maintaining an active stem-cell pool [
145]. Therefore, it is proposed that this mode of regulation has a general role in setting perennial flowering and in switching from monocarpic to polycarpic habits [
144].