Project Topics
Get Complete Project Material File(s) Now! »
Role of microRNAs
microRNAs are major regulators of gene expression in plants and their roles have been described in various processes from plant development to response to biotic and abiotic stresses but also in trans-kingdom communication between pathogens and their hosts (Sunkar et al., 2007; Weiberg et al., 2015; Couzigou and Combier, 2016). NGS analysis of small RNAs identify many microRNAs DE in various biological conditions suggesting a role for these microRNAs in these processes. However, functional validation is needed to test the functions of these genes, in particular by using mutants of these miRNAs and their target. In this chapter, I will present plant miRNAs for which a role has been biologically validated.
MicroRNAs in plant development
microRNAs regulate shoot, leaf and root developmental processes. Some microRNAs can act individually while other miRNAs regulate coordinately their targets during plant developmental processes.
Multiple microRNAs and their targets were shown to regulate leaf development. Leaf development mostly relies on the function of the shoot apical meristem (SAM). It begins with the initiation of leaf primordia then polarity establishment till the leaf acquired its final shape and size (Laufs et al., 1998; Barton, 2010). miR394 and its target LEAF CURLING RESPONSIVENESS (LCR); miR396/GROWTH REGULATING FACTORS (GRFs) and miR319/TEOSINTE BRANCHED1, CYCLOIDEA and PROLIFERATING CELL NUCLEAR ANTIGEN BINDING FACTOR (TCP) are involved in the control of SAM (Schoof et al., 2000; Efroni et al., 2008; Schommer et al., 2008; Liu et al., 2009; Baucher et al., 2013; Li et al., 2016a). Other microRNAs control leaf polarity like miR164 /CUP-SHAPED COTYLEDON2 (CUC2), miR165/miR166 /HOMEODOMAIN LEUCINE ZIPPER (HD-ZIP) and miR390/TAS3-tasiRNA (Peaucelle et al., 2007; Huijser and Schmid, 2011; Moon and Hake, 2011; Rubio-Somoza and Weigel, 2011; De Felippes et al., 2017).
An example of coordination of several miRNAs is the interplay between miR396 and miR319 and their targets in the control of cell proliferation and leaf shape in A. thaliana (Schommer et al., 2014), also with miR164 in the regulation of the shape and the size of the limbus (Rubio-Somoza and Weigel, 2011) and with miR156 in the switch from the juvenile to the adult phases of vegetative development (Rodriguez et al., 2016) (Figure 18). miR396 regulates leaf morphology by targeting GROWTH REGULATION FACTORS (GRF) transcription factor family (Liu et al., 2009; Baucher et al., 2013). GRFs are highly expressed in leaf meristematic tissue and positively regulate leaf size by promoting cell proliferation (Kim et al., 2003). Overexpression of miR396 induces the silencing of six GRF genes and leads to the formation of narrow leaves with a reduced size like the grf KO mutants (Horiguchi et al., 2005; Jeong and Byung, 2006). The expression of MIR396 is activated by TCP4 which is targeted by miR319 that is involved in leaf shape development (Palatnik et al., 2007; Schommer et al., 2014). A. thaliana transgenic plants overexpressing miR319 have increased leaf size, and a similar phenotype is observed in the tcp KO mutants (Schommer et al., 2008). Significant changes in TCP4 transcripts levels affect organ curvature. TCP4 directly activates the promoters of miR396 and miR164. miR164 targets CUC transcription factors whose activities contribute positively to the generation of leaf serrations. miR156 targets SPL transcription factors which promote the phase change from the juvenile to the adult phases of vegetative development and also to reproductive development; but they also influence leaf growth itself. The miR156, miR319 and miR164 networks are further interconnected by the protein-protein interactions of their targets. During the development of the younger leaves miR156 levels are high and therefore SPL levels are low, and TCP proteins form dimers with CUC proteins which in turn leads to smooth margins. Later on in development, when miR156 level goes down, SPL level increases and the TCP-CUC dimers are replaced by TCP-SPL dimers. The released CUC proteins dimerize and leaf serrations are formed.
The flowering stage, is regulated by an important number of miRNAs and their targets to synchronize flowering (Spanudakis and Jackson, 2014; Teotia and Tang, 2015; Samad et al.,
2017). We can cite for example, miR172/APETALA2-like (AP2) (Zhu and Helliwell, 2011; Teotia and Tang, 2015; Tripathi et al., 2018; Ó’Maoiléidigh et al., 2021), miR156/ SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) (Chuck et al., 2007; Efroni et al., 2008; Yamaguchi and Abe, 2012; Hong and Jackson, 2015; Teotia and Tang, 2015; Tripathi et al., 2018) and miR319/TCP (Schommer et al., 2008). As described above for leaf development, these microRNAs act coordinately to regulate transition from vegetative to flowering phase. miR156 family targets 11 of the 17 SPL genes. Among these genes, SPL3, 4, and 5 promote floral transition (Chuck et al., 2007; Xie et al., 2012; Hong and Jackson, 2015; Teotia and Tang, 2015). Tomato plants overexpressing SlymiR156a showed a delay in flowering and an extended juvenile phase (Zhang et al., 2011b). miR172 repress translation of AP2, a floral repressing transcription factor inducing defect in floral organ identity and mimicking the phenotype of the loss-of-function ap2 mutants (Chen, 2004; Teotia and Tang, 2015). Different studies highlighted the complex organization of miR172/AP2 and miR156/SPL module in stabilizing the floral state (Figure 19). In A. thaliana, miR156 and miR172 act together: in vegetative phase, miR156 maintains the juvenile phase by repressing SPL15 which inhibits floral transition. While in the flowering phase, TCP15 activates MIR172 expression that represses AP2 expression that induces the activation of floral transition (Teotia and Tang, 2015; Tripathi et al., 2018; Lian et al., 2021; Ó’Maoiléidigh et al., 2021).
Figure 19. Schematic representation of the interactions between miR172 and miR156 and their targets during flower transition; Crosstalk between miR156 and miR172 modules are shown along with the regulatory networks and feedback regulation of the target genes. miR156 is regulated by positive and negative feedback loop of SPL9 and SPL15, respectively; and positively regulated by AP2 and AGL15. miR172 is regulated by the positive feedback loop of TOE1/2 and negatively by AP2 through LUG and SEU. TOE1/2 repress the expression of SPL3/4/5 genes. SPL3 positively regulates the expression of TOE3. AP2 and SMZ repress their own expression and also of other miR172 target genes. AGL15, AGAMOUS LIKE15; AP2, APETALA2; LUG, LEUNIG; SEU, SEUSS; SMZ, SCHLAFMU ̈TZE; SNZ, SCHNARCHZAPFEN; SPL, SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE; TOE1-3, TARGETOF EAT1-3. (Teotia and Tang, 2015)
The root growth is sustained by the root apical meristem (RAM) which contains undifferentiated stem cells able to divide and differentiate to produce the different tissues of the root (Petricka et al., 2012). The function of miR160 (Mallory et al., 2005; Wang et al., 2005) and miR396 (Rodriguez et al., 2010; Bazin et al., 2013) has been established in RAM. It is important to mention that most miRNAs that are involved in root development are involved also in auxin and cytokinin hormone pathway. These two hormones are well known modulators of root formation and the cross-talk between them regulates RAM maintenance and lateral root emergence (Bishopp et al., 2011; Bielach et al., 2012). Thus, miR156 and its target SPL10 control root meristem activity and root-derived de novo shoot regeneration via cytokinin in A. thaliana (Barrera-Rojas et al., 2020). Arabidopsis plants overexpressing miR156a showed a reduced meristem size. The opposite root phenotype was observed in knock down miR156a expressing short tandem target mimic (STTM) that act as molecular sponge to sequester the microRNA and block its action to cleave the targets. Interestingly, CRISPR/Cas9-derived spl10-2 mutant (35-bp deletion in SPL10) displayed shorter meristems whereas rSPL10 lines that resist to miR156a cleavage showed the opposite phenotype suggesting that miR156 contributes to modulation of root meristem activity by targeting SPL10. Cytokinin controls root meristem size by activating the transcription factors of type-B ARABIDOPSIS THALIANA RESPONSE REGULATORS (ARRs) (Mason et al., 2004). ARR5 fused to the gene reporter GUS showed a strong expression in root explants of plants overexpressing miR156a. Conversely, weaker GUS staining was observed in rspl10 mutants resistant to cleavage. Moreover, medium supplement with different cytokinin presented an important regenerative capacity for controlling root explants, but it did not improve the low regenerative capacity of plants overexpressing miR156 neither for rspl10. All together, these results indicated that the meristem activity is regulated by miR156-SPL10 module probably through the reduction of cytokinin responses, via the modulation of ARR expression (Barrera-Rojas et al., 2020).
Moreover, miR393 (Figure 20) , miR390 (Chen et al., 2011; Lu et al., 2018), miR164 (Li et al., 2012; Geng et al., 2020), miR167 (Gutierrez et al., 2009), miR476 (Xu et al., 2021a) or miR847 (Wang and Guo, 2015) have been shown to be involved in the formation of lateral and adventitious roots. Several of these miRNAs regulate lateral root formation via Auxin Response Factor (ARFs). ARFs are key transcriptional factors regulators of auxin signaling that bind to auxin response elements (AuxREs) included in promoters of numerous auxin-responsive genes (Reviewed in Guilfoyle and Hagen, 2007; Chandler, 2016; Li et al., 2016b). miR390 induces the production of tasiRNAs that silence ARF2, ARF3 and ARF4 removing the repression of lateral root growth (Marin et al., 2010). miR160 is known to repress ARF17 (negative regulators of adventitious rooting) while miR167 repress the two targets ARF6 and ARF8 (positive regulators of adventitious rooting). A complex regulation and interaction between the three ARFs (ARF6, ARF8 and ARF17) and miR160/miR167 was observed using different grf mutant lines (Gutierrez et al., 2009). Thus, the balance between these two miRNAs control lateral root development (Couzigou and Combier, 2016).
Figure 20. Regulation of miR393 and its target in root development. A. thaliana lateral roots (A and E) Col-0 compared with mutants (B) tir1-1 and mutants overexpressing (C) miR393a, (D) miR393b, (F) TIR1 and (G) miR393-resistant form of TIR1 (mTIR1). Overexpressing (F) TIR1 and (G) mTIR1 displayed shorter primary roots, and more lateral roots while (B) tir1 and (C and D) miR393a/b overexpression displayed slightly longer primary root and fewer lateral roots, compared to wild type (A and E) Col0. TIR1, TRANSPORT INHIBITOR RESPONSE1. Bars = 10 mm (Chen et al., 2011)
MicroRNAs in plant response to abiotic stress
Plants are subjected to different environmental challenges such changes in temperature, soil water potential and nutrients. Induced phenotypic and physiological changes help the plant to adapt and survive in response to environmental changes (Figure 21) (Singh et al., 2020). Interestingly, some miRNAs respond to different abiotic stresses such as miR408 in response to cold, drought and copper stresses (Ma et al., 2015).
Under drought or salinity stresses, several miRNAs are differentially expressed and can positively or negatively regulate plant tolerance to drought stress (Song et al., 2013; Fang et al., 2014; Ma et al., 2015) or salinity stress (Jung and Kang, 2007; Song et al., 2013; Zhou et al., 2013; Iglesias et al., 2014; Kong et al., 2014). miR169 is a negative regulator of drought stress. In A. thaliana, plants overexpressing miR169 have an increase in susceptibility towards drought stress. Mutant overexpressing miR169 showed increased in size of the stomatal aperture that mimics the phenotype observed in miR169 target mutant nfya5 (NUCLEAR FACTOR Y SUBUNIT A5) (Li et al., 2008). miR394 and its target LEAF CURLING RESPONSIVENESS (LCR) are involved in plant responses to both drought and salinity stresses (Song et al., 2013). Using RT-PCR and GUS reporter fusion, miR394a and miR394b were showed to be induced under saline and drought stresses in A. thaliana. Conversely, plants overexpressing miR394b and lcr mutant showed a tolerance to drought stress. On the other hand, treatment with 100 mM NaCl showed a decrease in the germination of plants overexpressing miR394a and lcr seeds compared to wild type. These results suggest that both miR394 and LCR are critical for plant response to salt and drought stresses.
Table of contents :
1. Generalities
2. Plant-parasitic nematode
2.1 Ectoparasitic nematodes
2.2 Endoparasitic nematodes
2.3 Heteroderidae family
2.3.1 Cyst nematodes
2.3.2 Root-knot nematodes
2.3.2.1 M. incognita life cycle
2.3.2.2 RKN control strategies
3. Molecular mechanisms of plant-RKN susceptible interaction
3.1 RKN effectors
3.1.1 Effectors involved in the migration and cell wall degradation
3.1.2 Effectors involved in the suppression of defense reactions
3.1.3 Effectors involved in the formation of giant cells
3.2 RKN- induced feeding site
3.2.1 Morphology and ontogenesis of RKN-induced feeding site
3.2.2 Neighboring cells vs Giant cells
3.2.3 Analysis of gene expression in galls and giant cells
3.2.4 Key processes involved in formation of giant cells
3.2.4.1 Metabolism
3.2.4.2 Cell wall modification
3.2.4.3 Cell cycle regulation
3.2.4.4 Cytoskeleton reorganization
3.2.4.5 Defense response
3.2.4.6 Modulation of auxin and cytokinin phytohormones
4. MicroRNAs
4.1 Generalities
4.2 microRNAs and PTGS
4.2.1 Biogenesis and maturation of microRNAs
4.2.2 Mode of action of microRNAs
4.3 Role of microRNAs
4.3.1 MicroRNAs in plant development
4.3.2 MicroRNAs in plant response to abiotic stress
4.3.3 MicroRNAs in plant-microorganism interactions
