MicroRNAs As Potential Targets for Abiotic Stress Tolerance in Plants

18 Pages·2016·0.72 MB·English

by  Vinay Kumar

REVIEW
published:14June2016
doi:10.3389/fpls.2016.00817
MicroRNAs As Potential Targets for
Abiotic Stress Tolerance in Plants
VarshaShriram1,VinayKumar2*,RachayyaM.Devarumath3,TusharS.Khare2and
ShabirH.Wani4
1DepartmentofBotany,Prof.RamkrishnaMoreArts,CommerceandScienceCollege,SavitribaiPhulePuneUniversity,
Pune,India,2DepartmentofBiotechnology,ModernCollegeofArts,ScienceandCommerce,SavitribaiPhulePune
University,Pune,India,3MolecularBiologyandGeneticEngineeringSection,VasantdadaSugarInstitute,Pune,India,
4DivisionofGeneticsandPlantBreeding,FacultyofAgricultureWADURA,Sher-e-KashmirUniversityofAgriculturalSciences
andTechnology,Kashmir,India
The microRNAs (miRNAs) are small (20–24 nt) sized, non-coding, single stranded
riboregulator RNAs abundant in higher organisms. Recent findings have established
that plants assign miRNAs as critical post-transcriptional regulators of gene expression
in sequence-specific manner to respond to numerous abiotic stresses they face
during their growth cycle. These small RNAs regulate gene expression via translational
inhibition. Usually, stress induced miRNAs downregulate their target mRNAs, whereas,
their downregulation leads to accumulation and function of positive regulators. In the
past decade, investigations were mainly aimed to identify plant miRNAs, responsive
to individual or multiple environmental factors, profiling their expression patterns and
recognizingtheirrolesinstressresponsesandtolerance.AlteredexpressionsofmiRNAs
Editedby: implicatedinplantgrowthanddevelopmenthavebeenreportedinseveralplantspecies
NokwandaMakunga, subjected to abiotic stress conditions such as drought, salinity, extreme temperatures,
StellenboschUniversity,SouthAfrica
nutrient deprivation, and heavy metals. These findings indicate that miRNAs may hold
Reviewedby:
thekeyaspotentialtargetsforgeneticmanipulationstoengineerabioticstresstolerance
BiswapriyaBiswavasMisra,
UniversityofFlorida,USA in crop plants. This review is aimed to provide recent updates on plant miRNAs,
TarasP.Pasternak,
their biogenesis and functions, target prediction and identification, computational tools
InstituteofBiologyII,Germany
and databases available for plant miRNAs, and their roles in abiotic stress-responses
*Correspondence:
VinayKumar and adaptive mechanisms in major crop plants. Besides, the recent case studies
[email protected] for overexpressing the selected miRNAs for miRNA-mediated enhanced abiotic stress
toleranceoftransgenicplantshavebeendiscussed.
Specialtysection:
Thisarticlewassubmittedto
Keywords:abioticstress,microRNA,post-transcriptionalregulation,stress-responses,transgenics
PlantBiotechnology,
asectionofthejournal
FrontiersinPlantScience
INTRODUCTION
Received:21March2016
Accepted:25May2016 Plants being sessile organisms, persistently face adverse environmental perturbations termed as
Published:14June2016
abiotic stresses, most important being drought, soil salinity, extreme temperatures, and heavy
Citation: metals. Abiotic stresses have become a major challenge due to their widespread nature and the
ShriramV,KumarV,DevarumathRM, devastatingimpactsonplantgrowth,yieldsandthequalityofplantproduce.However,plantshave
KhareTSandWaniSH(2016)
developedintricatemechanismsforsensingandrespondingtoenvironmentalchanges(Wanietal.,
MicroRNAsAsPotentialTargetsfor
2016).
AbioticStressToleranceinPlants.
Front.PlantSci.7:817. To turn on protective mechanisms, plants trigger a network of genetic regulations including
doi:10.3389/fpls.2016.00817 alteredexpressionoflargeproportionofgenesbytranscriptionaland/ortranslationalregulations
FrontiersinPlantScience|www.frontiersin.org 1 June2016|Volume7|Article817
Shrirametal. PlantmiRNAsforStressTolerance
(Ku et al., 2015). Plants up-regulate the protective genes while of gene regulators, and these molecules are suggested to share
down-regulatingthenegativeregulators.Severalprotein-coding similar regulatory logistics besides participating in cooperative
genes have been recognized in recent years for controlling activitiesingeneregulatorynetworks(Linetal.,2012).
plant responses to abiotic stresses; however, our knowledge on ThediscoveryofmiRNAgenesinCaenorhabditiselegans(Lee
the regulatory mechanisms involved in this response are still etal.,1993)hasbeenfollowedbyasharpincreaseinidentification
limited and necessitates transformative tools to adapt crops of more and more plant miRNA families, making them a
to harsh environments (Zhang and Wang, 2015). These post- research hotspot (Cao et al. 2014; Supplementary Table S1 and
transcriptional regulations are pivotal for plants to restore and Supplementary Figure S1). A steady progress in investigations
re-establishtheircellularhomeostasisduringandrecoveryfrom involving miRNAs has led to a better understanding of
stressphases,respectively(Sunkaretal.,2012). transcriptional and post-transcriptional level gene-regulatory
RecentresearchindicatesthatplantsassignmiRNAsascritical mechanisms in plants (Sunkar et al., 2012; Zhang, 2015).
post-transcriptional gene-expression regulators to attenuate These tiny-sized non-coding molecules derived from stem-
plant growth and development under stress conditions, though loop structures are regarded as ubiquitous repressors of gene-
howthisisachievedatmolecularlevelsisyettobeunderstood expression as they fine-tune the target gene expression and
withgreaterdetails.ThemiRNAsrepresentawidespreadclassof degradeand/orinhibitproteinproductioninhighereukaryotes
small(20–24nt)endogenousRNAs(Zhang,2015)andregulate (Akdogan et al., 2015). It exemplifies the emerging view that
the gene expression via directing mRNA cleavage, translational miRNAs rival the proteins in regulatory importance (Mishra
repression, chromatin remodeling, and/or DNA methylation. et al., 2015). Nevertheless, this regulation is highly critical
Usually, stress-upregulated miRNAs down-regulate their target for all biological processes as well as stress-response and
mRNAs, whereas, their suppression leads to accumulation and acclimatization,thushasabigimpactonlifeprocessesofplants
functionofpositiveregulators(Chinnusamyetal.,2007).Several asindicatedbyseveralrecentinvestigations(ZhangandWang,
studies have confirmed that abiotic stress conditions induce 2015;LiandZhang,2016).MiRNAscontrolthegeneexpression
aberrant expressions of miRNAs in plants. High throughput viacausingepigeneticchangesbesidescontrollingtargetsatpost-
sequencing and computational approaches have been used in transcriptionallevel(Khraiweshetal.,2010;Wuetal.,2010).
recent year for identifying a large number of stress-related Recent findings affirmed that miRNAs play an array
miRNAs. These findings indicate that miRNAs might serve of important roles in plant growth, development and
as potential targets for genetic manipulations to engineer metabolism along with their involvement in abiotic stress
abiotic stress tolerance in plants. We summarize herein recent and pathogen responses (Yang C. et al., 2013; Xie et al., 2015).
updates on plant miRNAs, their biogenesis, target-genes, and MiRNAs have been reported as key regulators of plant root
their regulatory roles in abiotic stress-responses and adaptive development architecture via targeting AUXIN RESPONSE
mechanisms deciphered in major crop plants. Besides, the TRANSCRIPTION FACTOR (ARFs) (Khan et al., 2011). On
overexpression of selected miRNAs for miRNA-mediated plant similar lines, in a recent study, Ripoll et al. (2015) observed
stress tolerance has also been discussed. We have also tried to miRNA-regulated fruit growth in Arabidopsis. Besides, plant
shed light upon recent successes, current challenges and future miRNAs are also known to target genes involved in processes
directionsinthisfield. such as sulfur assimilation and ubiquitin-dependent protein
degradation (Bonnet et al., 2004). Since vegetative parts are
important in crop plants for harvesting reasons, therefore
PLANT miRNAs: TINY SIZE MAJOR ROLES control of plant apical dominance and vegetative growth is
significant. There is increasing evidence that miRNAs play
Production of abiotic stress tolerant crops necessitates better crucialroleinleaf-development,apicaldominanceandbiomass
understandingofgene-regulationmechanismsemployedbythe production, and targeting specific miRNAs is proving a novel
plantsinresponsetotheseenvironmentalcues.Researchfocused and potent strategy for improving plant growth, biomass and
on deciphering post-transcriptional regulations by non-protein cropyield(ZhangandWang,2015).
codingsmallRNAsthatconsistsblockingofspecificmessenger Computational predictions coupled with experimental
RNAs (mRNAs) or affecting epigenetic modifications at the approacheshaveledtotheconclusionthatmanyTFsincluding
transcriptional level has gained unprecedented attention. The MYBareindeedmiRNAtargets.Deepsequencinghasconfirmed
family members of these small ribonucleotide sequences are the critical roles miRNAs play in cotton ovule and/or fiber
represented by RNA species differing from each other on the development(Xieetal.,2015).Theyregulatekeygenesinvolved
basisoftheirsize,biogenesis,modeofaction,and/orregulatory in the floral induction and flower formation processes such
role (Mittal et al., 2016). MiRNAs are abundant in plants and as transition phases from juvenile to adult, initiation of floral-
areattributedformajorrolesinpost-transcriptionalregulations competenceandflowerdevelopment(HongandJackson,2015).
through base-pairing with complementary mRNA targets, Fruit and seed development held a distinct place in plant
particularlytranscriptionfactors(TFs)(LiandZhang,2016).In propagation and harvesting for defining crop yields, therefore,
plants, post transcriptional gene regulation involves miRNAs, the roles played by the miRNAs in development of fruits and
generatedbyDicer-like1(DCL1)frommiRNAprecursorsthat seeds is of great interest. Itaya et al. (2007) identified a large
are transcribed from miRNA genes (Mangrauthia et al., 2013). number of species-specific miRNAs from tomato (Solanum
Plant miRNAs along with TFs constitutes two major families lycopersicon) and hypothesized their significant roles in fruit
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Shrirametal. PlantmiRNAsforStressTolerance
development. Further, tissue-specific expression of numerous transcriptsthroughsequencecomplementarities(Voinnet,2009;
miRNAs was reported by Moxon et al. (2008) and the authors Khraiweshetal.,2010,2012).
proposed miRNA-regulated fleshy fruit development and The successful identification and binding of miRNA-AGO
ripening in S. lycopersicon. A number of miRNAs have been complex with specific mRNA-targets negatively regulate
identified for targeting metabolic pathways of grain-filing their expression via mRNA cleavage, decay, and/or post-
and nutrient-biosynthesis including carbohydrate and protein transcriptional repression (Jones-Rhoades et al., 2006). Though
metabolism, cellular transport and signal transduction besides there is clear evidence that the level as well as positions of
phytohormone signaling (Meng et al., 2013; Peng et al., 2013; complementarities between the mRNA targets and miRNAs
Zhang and Wang, 2015). MiRNAs have been attributed for defines the target selectivity, however, there is possibility of
vital functions in somatic embryogenesis in cotton (Gossypium involvement of other factors as well (Wang et al., 2015). Since
hirsutum) as revealed by the high-throughput and degradome all the miRNAs regulate plant growth and development as
sequencing(YangX.etal.,2013).Interestingly,decipheringthe well as stress-responses via targeting individual protein-coding
regulatory roles of miRNAs is not confined only to primary genes, therefore identification of these mRNA targets is the
metabolites.Recently,itwasrevealedthatcertainmiRNAstarget first and most important step in elucidating the functions of
transcriptsrelatedtosecondarymetabolismaswell(Bokeetal., miRNAs (Zhang and Wang, 2015). Once incorporated in RISC
2015; Bulgakov and Avramenko, 2015) and of these miR156, as discussed above, usually the mature miRNA regulate the
miR163,miR393,andmiR828havebeenascribedasinteresting expression of target genes via three mechanisms. It induces
toolsforregulatingsecondarymetabolisminArabidopsis. mRNA-slicing of specific target-gene and it is a main pathway
of miRNA action in plants (He et al., 2014), whereas second
pathway include the inhibition of translation of functional
PLANT miRNAs: THEIR BIOGENESIS, proteins through combining target gene’s mRNA and third
TARGETS AND MODE OF ACTION one incorporates silencing of multi-genes at transcriptional
level via chromosomal isochromatin (He et al., 2014). Owing
Much research, ever since the first report on plant miRNAs in to the fact that plant miRNAs represents the non-coding
2002(Reinhartetal.,2002)hasresultedinaconsiderablegainin regions of the genes, they act as an independent transcription
ourunderstandingoftheirorigin.Owingtotheubiquitousness unit. TFs are their major targets and have therefore critical
and diversity of plant miRNAs, it is apparent that most, if not roles in hormonal signal transduction, cellular metabolism,
all,biologicalprocessesinplantsatsomepointinvolvetheaction organ differentiation, floral patterning and reproduction,
of one or more miRNAs (Voinnet, 2009). Plant miRNAs share as well as in nutrient homeostasis and plant responses to
many similarities with their animal counterparts as revealed stresses (Sun, 2012; Hong and Jackson, 2015; Tripathi et al.,
by the recent genetic and biochemical studies focused on 2015).
decipheringtheirorigin,biogenesisandmodesofaction.These
smallbutfundamentalmoleculesaresynthesizedfromprimary
IDENTIFICATION AND TARGET
(pri)-miRNA transcripts, which are in turn transcribed usually
PREDICTION OF PLANT miRNAs:
by RNA pol II or pol III from the nuclear-encoded miRNA
(MIR)genes.Thepri-miRNAtranscriptsformdoublestranded COMPUTATIONAL TOOLS AND
stem-loop (imperfectly paired hairpin) structured precursor RESOURCES
RNAs(pre-miRNA).Inplants,apparentlyRNA-bindingprotein
DAWDLE(DDL)stabilizesthepri-miRNAsfortheirconversion To identify miRNAs in plants, both experimental as well as
to stem-loop pre-miRNAs in D-bodies. This reaction requires computational approaches have been used so far. Identification
a concentrated action and physical interaction of SERRATE of miRNAs in plants started in early 2000s with direct-cloning
(SE, a C2H2-zinc finger protein), a ds-RNA binding protein approaches (Llave et al., 2002; Reinhart et al., 2002). Though,
HYPONASTIC LEAVES1 (HYL1), DCL-1 and nuclear cap- it was not easy to identify large number of miRNAs, owing to
binding complex (CBC) (Fang and Spector, 2007; Gregory their multiple occurrences, small size and methylation status.
et al., 2008; Laubinger et al., 2008; Voinnet, 2009). The pre- But advancement in cloning methodologies and computational
miRNA hairpin precursor then gets converted into 20–22 nt algorithms resulted in a rapid increase in number of miRNAs
∗
miRNA::miRNA duplex, where miRNA acts as guide-strand identified in plants in last few years (Tripathi et al., 2015). In
∗
whereas miRNA as degraded strand by the actions of DCL1, recentpast,theadventofhighthroughput/deepsequencinghas
′
HYL1, and SE. This duplex then undergoes methylation at 3 really helped in an exponential growth in the number of plant
terminusbyHUAENHANCER1(HEN1)toprotectthemagainst miRNAs, not only identified but also functionally annotated
degradation by small RNA degrading exonuclease, SDN. The (Jagadeeswaran et al., 2010; Rosewick et al., 2013). This has
pre-miRNAs or the mature miRNAs are then exported from resulted into establishment of biological databases to act as
nucleustothecytoplasmbytheplantexportinproteinHASTY archives of miRNA-sequences and annotation such as miRBase
(Chaves et al., 2015). One strand of the duplex in cytoplasm (Tripathietal.,2015).
getsunwoundintoasinglestrandedmature-miRNAwhichthen Several computational tools and databases have been
gets incorporated into an ARGONAUTE protein, and directs developed in recent times to identify and predict the targets of
RNAinducedsilencingcomplex(RISC)bindingtocognatetarget miRNAsusingtraditionalnucleotidedatabasesaswellasdatasets
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Shrirametal. PlantmiRNAsforStressTolerance
generatedthroughdeep-sequencingapproaches,adetailedlistis identify hairpin excision regions and hairpin structure filtering
providedinTable1. forplants.Interestingly,itdoesnotneedthirdpartytools,rather,
The miRBase (http://www.mirbase.org/) represents a itusesagraphicuserinterfaceforinputandoutputofthedata,
comprehensive database with searchable online repository of andthedisplayofrecognizedmiRNAwithRNAseqreadsisdone
published miRNA sequences and associated annotation. This withthehelpofahairpindiagram.
databasewasstartedin2002withitsfirstreleaseincludedjust5 PlantMirnaTdevelopedbyRheeetal.(2015)isamiRNAand
miRNAsfromoneplantspecies(Arabidopsisthaliana),whereas mRNA integrated analysis system via utilizing the sequencing
itslatestversion(Release21,June2014)contains28,645entries dataeffectively.Itsmajorfeaturesincludeashortreadmapping
representing hairpin precursor miRNAs, expressing 35,828 tool,andanalgorithmwhichtakesintoconsiderationthemiRNA
mature miRNA products in 223 species including 73 plant expressionanddistributionintargetmRNAs(Rheeetal.,2015;
species with 7057 miRNA loci between them (Kozomara and https://sites.google.com/site/biohealthinformaticslab/resources).
Griffiths-Jones,2014). miTRATA (miRNA-Truncation and Tailing Analysis) is
miRNEST represents a wide-ranging database of animal, a web-tool for truncation and tailing analysis of miRNA,
′
plant, and virus miRNAs (Szczesniak et al., 2011, http:// and utilizes the miRBase (var. 21) and useful to analyze 3
mirnest.amu.edu.pl). It provides miRNA data on the basis of modifications of miRNAs (Patel et al. 2016). In recent years,
computationalpredictionsandhigh-throughputsequencing.It’s thoroughmiRNAbiogenesishasrevealedmorecomplexfeatures
current(miRNEST2.0)versioncontainsmiRNAsfrom22viruses and secondary structures of their precursors. Consequently,
and>270plantspecies.Interestingly,thisdatabasealsoincludes Eversetal.(2015)proposedafreelyavailablepublicaccesstool
degradomedataof2041entries(asonMarch16,2016). “miRA” (https://github.com/mhuttner/miRA) for identification
Plant miRNA database (PMRD, http://bioinformatics.cau. ofplant-miRNAprecursors,whichisadaptabletoheterogeneous
edu.cn/PMRD/) provides information about plant-miRNA and complex precursor populations. Interestingly, Meng et al.
sequencesaswellastheirtargets(Zhangetal.,2010).Itcontains (2016) developed a tool to predict the functions of plant
the sequence information, secondary structure, target genes, miRNAs on the basis of their functional similarity network
expression profiles and a genome browser. PMRD contains through application of transductive multi-label classification.
more than 8400 miRNA entries from >120 model plants as Transposable elements (TEs) have been recognized for their
well as major crops including Arabidopsis thaliana, rice (Oryza prominent roles in determining non-coding regions including
sativa),wheat(Triticumaestivum),soybean(Glycienemax),and miRNAsofthegenomes(Gimetal.,2014).Inordertodevelop
maize(Zeamays),besidesprovidingpredictedtarget-genesand a plant TE related miRNA database (PlanTE-MIR DB) was
interaction-site in the database. An updated version of PMRD proposedveryrecentlybyRLorenzettietal.(2016).Theauthors
hasbeenlaunchedasPNRD(PlantNon-codingRNADatabase) identified more than 150 miRNAs overlapping TEs in 10 plant
withgreaternumberofentriesandplantspeciesaswellasbroad genomes.Thispubicdatabaseishostedathttp://bioinfo-tool.cp.
spectrum RNA types (Yi et al., 2015). There are total 25,739 utfpr.edu.br/plantemirdb/.
entriesofnon-codingRNA(ncRNA)includinglncRNAs,tRNA, It is noteworthy to mention here that in spite of numerous
rRNA, tasiRNA, snRNA, and snoRNA from around 150 plant computational tools to identify plant-miRNAs and their target
species,especiallyfoodcrops.Itoffersvarioussearchandanalysis prediction, there is a lack of curated databases of stress
tools for the user such as ncRNA keyword search, for example related miRNAs. Zhang et al. (2013) advocated for a need
IDsearch,targetsearch,andtoolkitsforpredictingonlinenovel to construct a cohesive database system for data deposit and
miRNAs and for calculating coding potential along with the further applications of plant abiotic stress related miRNAs and
availability of BLAST tools (http://structuralbiology.cau.edu.cn/ accordinglydevelopedPASmiR,whichallowstheuserstoretrieve
PNRD/index.php). miRNA-stressregulatoryentriesbykeyword-searchsuchasplant
PMTED (Plant miRNA Target Expression Database, http:// speciesandtypeofstress(Zhangetal.,2013).Similarly,Remita
pmted.agrinome.org/) is a plant-specific miRNA database, et al. (2015) proposed a web-based server/database “WMP”
useful to study miRNA functions by inferring their target dedicated to wheat miRNAs, particularly stress-responsive
gene expression profiles among the large amount of existing miRNAsandisavailableat:http://wheat.bioinfo.uqam.ca.
microarray data. It contains tools to search miRNA targets,
retrieve expression data and the user can find out differentially
expressedgenes(Sunetal.,2013). miRNAs INVOLVED IN PLANT
The TAPIR (target prediction for plant miRNAs) webserver ABIOTIC-STRESS RESPONSES
(http://bioinformatics.psb.ugent.be/webtools/tapir/) predicts
targets for plant miRNAs, with two available modes, the first Through adaptive evolution processes, plants have developed
“Fast” mode using the FASTA search engine while the second incredibleabilitiestorespondandadapttochallengingexternal
“Precise” mode using the RNA-hybrid search engine (Bonnet conditions. The cellular and molecular responses of plants
etal.,2010). to these abiotic stresses are intricate and received great
miRPlant is an integrated tool for identification of plant attention of plant breeders and biotechnologists in recent
miRNA from RNA sequencing data (An et al., 2014, http: past to decipher them. Plants deploy multitude abiotic stress
//www.australianprostatecentre.org/research/software/mirplant). responsive mechanisms involving regulation of expression of
miRPlant works on the strategies specifically developed to stress-responsivegenestosenseandrespondtoexternalstimuli.
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TABLE1|AsummarizedlistofmajortoolsavailableforplantmiRNAs,theirtargetidentification/predictionandrepositories.
Nameofthe Description Weblink References
database/resource/
repository/tool
TAPIR TargetpredictionforPlantmiRs http://bioinformatics.psb.ugent.be/webtools/tapir/ Bonnetetal.,2010
miRTarBase TheexperimentallyvalidatedmiR-targetinteractions http://mirtarbase.mbc.nctu.edu.tw/index.php Hsuetal.,2011
database
PMRD PlantmiRNADatabase http://bioinformatics.cau.edu.cn/PMRD/ Zhangetal.,2010
miRanalyzer miRdetectionandanalysistoolfornext-generation http://bioinfo5.ugr.es/miRanalyzer/miRanalyzer.php Hackenbergetal.,2011
sequencingexperiments
PmiRKB PlantmiRKnowledgeBase. http://bis.zju.edu.cn/pmirkb/ Mengetal.,2011
Fourmajorfunctionalmodules,SNPs,Pri-miRs,
MiR-TarandSelf-reg,areprovided
miRDeep-P Acomputationaltoolforanalyzingthe http://faculty.virginia.edu/lilab/miRDP/ YangandLi,2011
miRtranscriptomeinplants
C-mii AtoolforplantmiRandtargetidentification http://www.biotec.or.th/isl/c-mii Numnarketal.,2012
Semirna SearchingforplantmiRNAsusingtargetsequences http://www.bioinfocabd.upo.es/semirna/ Muñoz-Méridaetal.,
2012
mirTool Acomprehensivewebserverprovidingdetailed http://centre.bioinformatics.zj.cn/mirtools/ Wuetal.,2013
annotationinformationforknownmiRsandpredicting
novelmiRsthathavenotbeencharacterizedbefore
PASmiR Aliterature-curateddatabaseformiRmolecular Zhangetal.,2013
regulationinplantresponsetoabioticstress
miRBase SearchabledatabaseofpublishedmiRsequencesand http://www.mirbase.org Kozomaraand
annotation Griffiths-Jones,2014
miRPlant AnIntegratedToolforIdentificationofPlantMiRfrom http://www.australianprostatecentre.org/research/ Anetal.(2014)
RNASequencingData software/mirplant
MTide AnintegratedtoolfortheidentificationofmiR-target http://bis.zju.edu.cn/MTide/ Zhangetal.,2014b
interactioninplants
PNRD ItisanupdatedversionofPMRD http://structuralbiology.cau.edu.cn/PNRD/index.php Yietal.,2015
PlantMirnaT AmiRNA-mRNAintegratedanalysissystem https://sites.google.com/site/biohealthinformaticslab/ Rheeetal.,2015
resources
miRA PlantmiRNAidentificationtoolespeciallyfororganisms https://github.com/mhuttner/miRA Eversetal.,2015
withoutexistingmiRNAannotation.Itisalsousefulfor
identifyingspecies-specificmiRNAs
miPEPs MiRNAsEncodePeptidesisatoolforfunctional Couzigouetal.,2015
analysisofplantmiRNAfamilymembers
sRNAtoolbox Asetoftoolsforexpressionprofilingandanalysisof http://bioinfo5.ugr.es/srnatoolbox Ruedaetal.,2015
sRNAbenchresults
miRge Afastmultiplexedmethodofprocessing http://atlas.pathology.jhu.edu/baras/miRge.html. Barasetal.,2015
sRNA-sequencedatatodeterminemiRNAentropy
andidentifydifferentialproductionofmiRNAisomiRs
BioVLAB-MMIA-NGS MiRNAandmRNAintegratedanalysisusing http://epigenomics.snu.ac.kr/biovlab_mmia_ngs/ Chaeetal.,2015
high-throughputsequencingdatacoupledwith
bioinformaticstools.
DMD AdietarymiRNAdatabasefrom15dietaryplantand http://sbbi.unl.edu/dmd/ Chiangetal.,2015
animalspecies
WMP DatabaseforabioticstressresponsivemiRNAsin http://wheat.bioinfo.uqam.ca Remitaetal.,2015
wheat
miTRATA AtoolformiRNAtruncationandtailinganalysis https://wasabi.dbi.udel.edu/~apps/ta/ Pateletal.,2016
MFSN AtoolforpredictionofplantmiRNAfunctionsbasedon Mengetal.,2016
functionalsimilaritynetwork(MFSN)through
applicationoftransductivemulti-labelclassification
(TRAM)totheMFSN
PlanTE-MIR Databasefortransposableelement-relatedplant http://bioinfo-tool.cp.utfpr.edu.br/plantemirdb/ RLorenzettietal.,2016
microRNAs
P-SAMS APlantSmallRNAMakerSite(P-SAMS)isawebtool http://p-sams.carringtonlab.org Fahlgrenetal.,2016
forartificialmiRNAsandsynthetictrans-actingsmall
interferingRNAs
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Investigations in recent times have shown that different abiotic In rice, Zhao et al. (2007) reported that miR169g and
stress conditions induce aberrant expression of thousands of miR393 are strongly upregulated and transiently induced,
protein-coding genes in various plant species (Kumar et al., respectively by drought. In another study, Zhou et al. (2010)
2009; Zeller et al., 2009; Turan and Tripathy, 2013; Khare used microarray approach for identify and analyze miRNAs
et al., 2015). Many of these stress-induced, often individual, in drought stressed rice at developmental stages ranging
genes for example proline biosynthetic pathway gene P5CS in from tillering to inflorescence formation. Overall, drought
rice (Kumar et al., 2010), methionine sulfoxide reductase gene conditions induced differential expressions of 30 miRNAs, out
(MSRB7) in Arabidopsis (Lee et al., 2014) and transcription of which 19 were newly reported drought-regulated miRNAs
factor TaERF3 in wheat (Rong et al., 2014) were overexpressed from Arabidopsis. Sixteen miRNAs (miR156, miR159, miR168,
to produce respective transgenic plants to confer enhanced miR170, miR171, miR172, miR319, miR396, miR397, miR408,
toleranceagainstsingularabioticstress.However,theregulatory miR529, miR896, miR1030, miR1035, miR1050, miR1088, and
mechanismsoftheseprotein-codinggenesarelargelyunknown miR1126)weredownregulatedbydroughtstresswhileitinduced
(Sun, 2012), in this regard, the miRNAs may prove extremely the overexpression of 14 miRNAs (miR159, miR169, miR171,
importantindecipheringthesegene-regulatorymechanismsand miR319, miR395, miR474, miR845, miR851, miR854, miR896,
thestress-responses.Though,largenumbersofmiRNA-families miR901, miR903, miR1026, and miR1125). Notably, miR171,
are conserved in flowering plants, however, recent updates miR319, and miR896 were identified in both (overexpressed
indicated that miRNAs may be species-, physiological stage or and repressed) groups (Zhou et al., 2010). Predicted targets
event-,organortissue-,andstress-specific(Valdes-Lopezetal., of the differentially regulated miRNAs were largely TFs (Zhou
2010;Sun,2012). et al., 2010). Similarly, drought stress induced expression was
Several miRNAs have been recognized as abiotic stress- observedin438miRNAsagainst205undercontrolledconditions
regulated in important crops and/or model plants under soil in Triticum turgidum leaf and root tissues, and 13 miRNAs
salinity(Gaoetal.,2011),nutrientdeficiency(Liangetal.,2015), weredifferentiallyregulatedbydroughtconditions(Kantaretal.,
UV-B radiation (Casadevall et al., 2013), heat (Goswami et al., 2010). Aberrant miRNA expression was observed in soybean
2014), and metal stress (Yang and Chen, 2013; Gupta et al., cultivars subjected to drought conditions, with upregulation
2014). However, this stress-regulated miRNA expression does of miR166-5p, miR169-3p, miR1513c, miR397ab, and miR-
not essentially confirm that the miRNA is involved in plant’s seq13insensitivecultivarsbutdownregulationintheirtolerant
adaptation to stress conditions. Zhang et al. (2013) observed counterparts(Kulcheskietal.,2011).
differential expression of as many as 1062 miRNAs in 41 plant Recently in sugarcane, the expression pattern of miRNA
species under 35 different types of abiotic stresses. Table2 was observed to be dependent on the species, type of stress,
summarizes recent data on the altered miRNA expression in tissue (seedlings, leaves, spikelets, and root) growth condition
majorcropplantsinresponsetoabioticstresses. (field, greenhouse, hydroponic culture system). MiR396 and
miR171 were differentially expressed in the most of the cases
(Gentileetal.,2015).Interestingly,overexpressionofosa-miR393
Drought and Salinity Stress resultedintoenhancedsalttolerance,suggestingtheirregulatory
Owingtothewidespreadchallengesthesetwostress-factorsput role in salinity tolerance in Arabidopsis (Gao et al., 2011).
on plant growth and yield, drought, and salinity stresses have In similar vein, Sun et al. (2015) identified 49 known and
received maximum attention of plant scientists to study their 22 novel salt stress-responsive miRNAs in radish (Raphanus
deleterious effects and decrypting the responsive-mechanisms sativus)andinterestinglythetargetpredictionanalysisrevealed
adopted by the plants. Over the past two decades, researchers the implication of the target genes in signaling, regulating ion-
investigated and identified number of genes to improve stress homeostasis besides modulating the decreased plant growth
tolerance of transgenics overexpressing those genes. However, undersaltstress.
ononehandtheresultanttransgenicplantsshowedinadequate
improvement largely because the complex genetic interactions Extreme Temperatures
and on the other hand their regulation underlying plant stress Plants often face extreme temperatures due to geographical
tolerance are not completely understood (Sunkar et al., 2012). conditions and seasonal variations, which negatively affects the
Non-coding RNAs including miRNAs however are turning to plant growth and productivity and plants adjust their gene
beapotentplatformtounderstandanddecodestress-responsive expression patterns at post-transcriptional levels in response
transcriptionalandpost-transcriptiongeneregulationsinplants. to these inconstant external temperatures. Numerous chilling
A number of drought stress-responsive miRNAs were or heat responsive miRNAs have been detected in plant
identified via screening of small RNAs library isolated from species (Cao et al., 2014). Wheat seedlings of heat tolerant
◦
Arabidopsis (Liu et al., 2008), rice (Zhou et al., 2010), and and susceptible cultivars when exposed to heat stress (40 C)
sugarcane(Gentileetal.,2015)andthecountiseverincreasing. exhibited differential miRNA gene expression with increased
Liuetal.(2008)evidencedatotalof14stress-induciblemiRNAs expression in many of them, where expression was suppressed
inArabidopsis,outofthem10wereNaCl-regulated,4drought- in few under heat stress (Xin et al., 2011). Eighteen cold-
◦
regulated and 10 were cold (4 C)-regulated, and miR168, responsive miRNAs were identified by Lv et al. (2010) in
miR171,andmiR396 (withTFsaspredictedtargets)responded rice with most of them being downregulated by the cold
◦
toallofthestresses. stress (4 C) and the authors hypothesized the miRNAs as
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Shrirametal. PlantmiRNAsforStressTolerance
TABLE2|DifferentialexpressionpatternsofmiRNAsreportedunderdifferentabioticstresses(S-salinity;D-drought;HT-hightemperature;C-cold;
Cd-cadmium;As-arsenic)inmajorcrops-Oryzasativa,Triticumaestivum,Saccharumsp.,Glycinemax,andHordeumvulgare.
miRNA Crop Response Validated/PutativeTarget
S D HT C Cd As
miR156 O.sativa SPLTFs
T.aestivum SPL
Saccharumsp. SPL5
G.max SPL
miR159 O.sativa MYBfamilyTFs
T.aestivum MYBdomainprotein
Saccharumsp. GA-Myb
miR160 O.sativa ARF
T.aestivum ARF
H.vulgare ARF13,ARF17
miR162a O.sativa DCL1
miR164 T.aestivum NACdomaincontainingprotein
miR166 O.sativa STARTdomaincontainingprotein,HD-ZipTFs
T.aestivum HD-Zipprotein
Saccharumsp. ClassIIIHD-Zipprotein4
H.vulgare HD-ZipTFs
miR167 O.sativa ARF
T.aestivum ARF
Saccharumsp. ARF17
G.max ARF
H.vulgare ARF8
miR168 O.sativa AGO
T.aestivum AGO
Saccharumsp. AGO1
H.vulgare AGO1
miR169 T.aestivum CBF-B/NF-YAfamilyprotein
Saccharumsp. HAP12-CCAAT-boxTFcomplex
O.sativa NF-YAorHAP2TFs
miR170 O.sativa Scarecrow-likeTF,ACP1
miR171 O.sativa Scarecrow-likeTF
T.aestivum Scarecrow-likeTF
Saccharumsp. SCL1
G.max Ubiquitincarrierprotein,Fe-SOD1
H.vulgare ProteinFAN
miR172 O.sativa APETALA2,bZIPTFfamilyprotein
T.aestivum APETALA2-like
Saccharumsp. APETALA2-like
miR319 O.sativa TCPfamilyTF21
miR390 O.sativa LRR-RLK
(Continued)
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Shrirametal. PlantmiRNAsforStressTolerance
TABLE2|Continued
miRNA Crop Response Validated/PutativeTarget
S D HT C Cd As
miR393 O.sativa TIR1,AFB2,AFB3,F-boxdomain,LRRcontaining
protein/MYBfamilyTF
miR394 O.sativa F-boxdomaincontainingprotein
miR395 T.aestivum ATPsulfurylase
G.max ATPsulfurylase,Lowaffinitysulfatetransporter
miR396 O.sativa GRFTFs,rhodenase-likeproteins,kinesin-likeproteinB
T.aestivum GRF3
Saccharumsp. GRF1
G.max CytochromeP450monooxygenase,NADH-ubiquinone
oxidoreductasechain4
miR397 Saccharumsp. LAC-10precursor
O.sativa LACs
miR398 Saccharumsp. Seleniumbindingprotein
miR408 O.sativa SPX,BCP
miR444 O.sativa MADS-boxTFs
H.vulgare MIKC-typeMADS-boxTFs
miR474 O.sativa PPR,proteinkinase,kinesin,leucine-richrepeat
miR482 T.aestivum Transmembraneprotongradientregulation
miR528 O.sativa IAR1,CBP/Plastocyanin-likedomaincontainingprotein
O.sativa OsDCL1
Saccharumsp. PutativeLAC
miR529 O.sativa SBP-boxgenefamily
miR530- O.sativa Hairpin-inducedprotein1domaincontainingprotein
3p
miR809 O.sativa Glutaredoxin2,putative,PPRrepeatcontainingprotein
miR827 T.aestivum NLA
miR1318 O.sativa CalciumbindingproteinsorCalciumATPases
miR1428 O.sativa Cytochromec
miR1432 O.sativa CalciumbindingproteinsorCalciumATPases
miR1884 O.sativa AAAATPase
miR2871 O.sativa GTfamilyprotein
miR5049 T.aestivum Wpk4proteinkinase
miR5175 H.vulgare ACC-likeoxidase
InthetabledifferentialexpressionpatternofmiRNAsunderdifferentabioticstressenvironments(salt,drought,hightemperature,cold,cadmium,andarsenic)wasmarkedasinduced
(greenbox)orrepressed(redbox)expression.References-Oryzasativa:Dingetal.(2011),Barrera-Figueroaetal.(2012),Zhouetal.(2010),Xiaetal.(2012),LiuandZhang(2012);
Triticumaestivum:Xinetal.(2010),Pandeyetal.(2014),Akdoganetal.(2015);Saccharam:Bottinoetal.(2013),Gentileetal.(2015);Glycinemax:Lietal.(2011);Hordiumvulgare:
Dengetal.(2015),Hackenbergetal.(2015),Kruszkaetal.(2014).
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Shrirametal. PlantmiRNAsforStressTolerance
ubiquitous regulators in rice. On the other hand, cold stress Heavy Metals (HMs)
brought a sharp increase in the expression of miR812q in rice Though heavy metals naturally occur in earth’s crust, but
plants at the beginning of reproductive phase (Jeong et al., anthropogenic and industrial activities has led to severe HM
2011).Theseobservationsholdmerit,sincemiR812qoriginates pollution in vast areas under cultivation. HM toxicity is one of
from a sequence-diverged region, has unique sequence, and themajorabioticstressesleadingtohazardouseffectsviaaltering
unlike other family members, is cold-responsive (Jeong et al., thephysiologicalandmetabolicprocessesthatnegativelyaffects
2011; Jeong and Green, 2013). Cold stress induced miR812q growth, development and productivity of crops (Rascio and
targetsCIPK10anddownregulatesthelater,interestingly,CIPK Navari-Izzo, 2011; Gupta et al., 2014). HMs can be categorized
transcripts are known abiotic stress tolerance mediators in into two groups, essential and non-essential HMs. Copper
calcium dependent CBL-CIPK signaling pathway (Jeong and (Cu), iron (Fe), zinc (Zn), and manganese (Mn) which at low
Green,2013). concentrations plays vital roles in enzymatic and biochemical
Such differential expression may be due to species-specific reactionsinplantcell,howeveraretoxicathigherconcentrations
role of miRNAs in cold response (Jeong and Green, 2013). (RascioandNavari-Izzo,2011;Gielenetal.,2012).Whereasnon-
In a recent study by Zhang et al. (2014a), high-throughput essentialmetalsconsistsofcadmium(Cd),aluminum(Al),and
sequencinganalysesrevealedcoldstress-inducedupregulationof mercury(Hg)andaretoxictoplantsevenatlowconcentrations
31anddownregulationof43miRNAsintea(Camelliasinensis) (Gielen et al., 2012). Though cobalt (Co) was historically not
plants and authors identified a large number of target genes consideredasvitalforplantgrowthanddevelopment,however,
using degradome sequencing. Functional analysis of miRNA itisnowclassifiedasessentialmicronutrient.Coplayssignificant
targetsreaffirmedtheircriticalrolesinstressresponse.Similarly, rolesingrowthanddevelopmentofplantbuds,leaf-discsstems
Cao et al. (2014) constructed small-RNA and degradome andcoleoptile,besidesassistinginCO absorptionbytheplants
2
libraries from wild tomato (Solanum habrochaites), and out (Diez, 2014). The HM entry to human food-chain through
of these, 192 miRNAs showed increased expression, while contaminated crops is a serious human health threat (Gupta
expression was decreased in 205 miRNAs. Besides, authors et al., 2014). Recent investigations have established miRNA-
also predicted the miRNA targets and few of the targets mediatedtranscriptionalandpost-transcriptionalregulationsof
were validated using qRT-PCR. Further, most of the target gene expression via base-pairing with their target mRNAs in
genes were reported to have positive role in chilling response plantssubjectedtoHM-stress(YangandChen,2013).
as revealed by their functional analysis via regulating the
expression anti-stress proteins and antioxidants (Cao et al., Cd-Toxicity
2014). Cd accumulation by plants results into chlorosis, wilting,
growth reduction and cell death, damage to proteins and
Hypoxia and Oxidative Stress induces oxidative stress (Zhao et al., 2012). Cd affects crop
Oxygenisextremelyimportantforaerobicorganismsincluding productivity and its exposure predominantly through food
plants. Hypoxia or low-oxygen stress usually results from but contamination pose serious risks to human health (Rizwan
not limited to the water logging conditions in the rhizospheric et al., 2016). Steady progress has been made in recent past
environments of plants. It eventually leads to diminished in investigating the molecular mechanisms involved in Cd
oxygenavailabilityandthusseverelyaffectingthemitochondrial toxicityandtolerance/adaptivemechanismsinplants.Aberrant
respiration (Agarwal and Grover, 2006). However, hypoxia miRNA expression in stressed plant cells and tissues indicated
also holds significance in signaling. It induces a metabolic involvementofmiRNAsinCd-responses.
shift from aerobic respiration to anaerobic fermentation Huangetal.(2010)documentedelevatedexpressionofbna-
and significant changes in transcriptome (Moldovan et al., miR393 in leaves, bna-miR156a, bna-miR167a/c in roots and
2010). Recent studies suggested a number of miRNAs as leaves, bna-miR164b and bna-miR394a/b/c in all tissues of
hypoxia-responsive and advocate them to be critical post- rapeseed (Brassica napus) upon Cd exposure with concomitant
transcriptional modulators (Licausi et al., 2011; Zhai et al., downregulationofmiR160.Theauthorsinanattempttopredict
2013). In Arabidopsis roots, Moldovan et al. (2010) reported thetargetsgeneratedtransgenicB.napuslinesbyoverexpressing
enhanced expression levels of miR156g, miR157d, miR158a, miR395 and confirmed its targets to be sulfate transporters
miR159a, miR172a,b, miR391, and miR775 by low oxygen (particularlySULTR2;1)andATPsufurylases(APS)(Huangetal.,
stress. 2010).SULTR2;1isvitalforitsroleinsulfateremobilizationfrom
In Arabidopsis, Sunkar et al. (2006) revealed repression of mature to younger leaves while APS catalyzes the first step in
miR398 and the upregulation of superoxide dismutase (SOD) thesulfurassimilation(Liangetal.,2010).Similarly,Zhouetal.
proteins under oxidative stress. A study of H O -regulated (2012a)reportedCdinduceddifferentialexpressionofnumerous
2 2
miRNAs from rice seedlings indicated that expression of some conservedandnon-conservedmiRNAsinB.napusroots.Using
miRNAs is regulated in response to oxidative stress; results deep-sequencing analysis, authors tried to understand how Cd
revealed that out of seven H O -responsive miRNAs i.e., regulates genome-wide regulation of miRNA expression and
2 2
miRNAs that were expressed differentially in H O -treated their targets, in all 84 miRNAs including conserved and non-
2 2
and control samples; miR169, miR397, miR827, and miR1425 conserved families were identified with most of the miRNAs
were upregulated while miR528 was downregulated by H O - exhibitingdifferentialexpressioninshoots/rootsunderCdstress
2 2
treatments(Lietal.,2010). (Zhouetal.,2012a).Inrice,usingmicroarrayassay,Dingetal.
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Shrirametal. PlantmiRNAsforStressTolerance
(2011)identified19Cd-responsivemiRNAsfromriceseedlings, miRNAs in common bean (Phaselous vulgaris) and a total
andtheirtargetswerepredictedastranscriptionfactorsandstress of 37 miRNAs displayed differential expression under abiotic
related proteins. In an attempt to decipher the Cd-responsive stresses including Mn. Out of these, eleven miRNAs were
miRNAs and their mediated gene regulatory networks at the induced whereas another eleven were inhibited under Mn
transcriptome level, Xu et al. (2013) constructed small RNA stress. miR1508, miR1515, miR1510/2110, and miR1532 were
librariesfromradish(Raphanussativus)withorwithoutexposed characterizedasMn-responsiveandtheirtargetswerepredicted
to Cd stress. They identified 15 known and 8 novel miRNA as calcium-dependent protein kinase, heat shock proteins,
familieswithnotabledifferentialexpressioninCd-stressedplants nucleotide-binding site leucine rich repeat resistance like
andtheirtargetswerethenpredictedusingdegradomeanalysis. proteinsandreceptorkinaseprotein,respectively(Valdes-Lopez
Authors postulated the target genes of Cd-responsive miRNAs etal.,2010).
as phytochelatin-synthase-1, and iron and ABC transporter
proteins(Xuetal.,2013).
As-Toxicity
Arsenic is a class-I carcinogen, widely distributed metalloid
Hg-Toxicity
Hg is a highly toxic metal and its ionic form (Hg2+) is presentintheearth’scrustandenforcesseverethreatstohuman
health via entering the food chain through contaminated food
most prevalent in soil and bioavailable form for plants which
crops (Srivastava et al., 2011). Investigations have shown that
are readily taken up through root and aerial part of the
arsenitetoxicityinplanttissuesleadstoreducedphotosynthesis
plants. It causes toxicity response such as stunted growth,
rate,perturbedcarbohydratemetabolism,besides,generationof
loss of cell shape, vascular abnormality, reduced chlorophyll
ROS, and lipid peroxidation (Jha and Dubey, 2004; Requejo
contentandreactiveoxygenspecies(ROS)accumulation(Chen
and Tena, 2005). However, little is known about the miRNA-
and Yang, 2012). Differential miRNA expressions have been
mediated post-transcriptional regulatory mechanisms involved
reported as a response to Hg exposure in various plant species,
inAsresponsesandtoleranceinplants.Therefore,itisinteresting
for instance, Zhou et al. (2012b) through deep sequencing
to investigate and understand regulatory roles miRNAs play
approach observed that numerous conserved as well as non-
in response to As-stress in plants. In rice, Yu et al. (2012)
conservedmiRNAsweredifferentiallyexpressedinbarrel-clover
identified36newmiRNAsresponsivetoAsandtheyareinvolved
(Medicago truncatula) seedlings subjected to Hg stress. A total
regulating gene expression in transportation, signaling and
of 130 targets for 58 known miRNA families comprising both
metabolism,thesefindingleadstojasomonicacid(JA)andlipid
conservedaswellasnon-conserved,besides,37targetsfornovel
metabolisminresponsetoAs-stress.Inaddition,LiuandZhang
M. truncatula-specific candidate miRNAs were identified using
(2012)reported67As-responsivemiRNAsfromindicariceroots
degradomesequencing(Zhouetal.,2012b).Authorsattributed
and showed that miR408, miR528, and miR397b had increased
thedifferentialexpressionofidentifiedmiRNAsandtheirtargets
expression,howevermiR1316andmiR390wererepressedunder
toHgstress.
As-stress.SincetheroleofJAinAsstressperceptionisestablished
Anunfortunateconsequenceofabioticstressesistheoxidative
(Srivastava et al., 2009), a few miRNAs have been identified
burst mediated through the excessive ROS generation in plant
to affect JA biosynthesis via targeting the transcription factor
tissuesandcells(Khareetal.,2015).Plantsdeployantioxidative
involvedtherein(Guptaetal.,2014),forinstance,Srivastavaetal.
armory with enzymatic and non-enzymatic components to
(2012)hasreportedthatmiR319hasbeenfoundtoberesponsive
detoxify or scavenge these ROS in a systematic way (Khare
toAsstress.ThisindicatesthecriticalrolesofplantmiRNAsin
et al., 2015). Recent reports confirm the role of miRNAs
regulatingJAbiosynthesisandwhichmightinturnbeinvolved
as post-transcriptional regulators of metal induced oxidative
inmetaltoxicityresponsesinplants.
stress and responsive antioxidation. It is exemplified by Hg-
induced enhanced activity of superoxide dismutase (SOD)
in alfalfa (Medicago sativa, Zhou et al., 2008). Further, the Al-Toxicity
overexpressionofmiR398-resistanttranscriptofCSD2(Cu/Zn- Al is a major toxicity factor, negatively affecting the crop
SOD 2) resulted into more CSD2 transcripts and enhanced production on 30–40% of the world’s arable land. It binds with
heavymetaltoleranceintransgenicArabidopsisovertheirnon- COOH and PO groups present on the root cell wall, which
4
transformed counterparts (Sunkar et al., 2006), suggesting the leads to structural changes, inhibition of cell wall expansion
apparentinvolvementofmiR398inHg-inducedresponses. anddiminishedrootgrowth(Maetal.,2004).Thisreducesthe
uptakeofmineralsnutrientandwater.Italsoaffectsphysiological
Mn-Toxicity processes like callose deposition, imbalance cytoplasmic Ca2+
Manganese acts as inorganic catalyst, while Mn and Fe and induce oxidative stress (Silva, 2012). In recent years, role
are important components for many enzymatic reactions. of miRNA using high-throughput genome sequencing, Chen
Comparably lesser research has been carried out to identify et al. (2012) reported altered expression of 23 miRNAs in M.
the Mn-toxicity regulated miRNAs, their targets and detailed truncatula seedlings subjected to Al-stress. Subsequently using
post-transcriptionalregulatorymechanismsinplantsinresponse same methodology Zeng et al. (2012) identified 30 miRNAs in
to Mn stress (Valdes-Lopez et al., 2010; Gupta et al., 2014). Al-treatedsoybean,andinterestingly,inthisstudymiR396 and
Valdes-Lopezetal.(2010)usedmiRNAmacroarrayhybridization miR390 were upregulated unlike earlier observations by Chen
approach coupled with qRT-PCR to identify Mn-responsive etal.(2012),indicatingspecies-specificnatureofthesemiRNAs.
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