MicroRNAs (miRNAs) are small non-coding RNA molecules involved in post-transcriptional gene silencing. They are 21-24 nucleotides long and regulate gene expression by binding to target mRNAs. The first miRNA, lin-4, was discovered in C. elegans in 1993. Currently over 24,000 miRNAs have been discovered across species. MiRNAs are important regulators of gene expression under stress conditions in plants. For example, miR399 is strongly induced during phosphate starvation and targets the PHO2 gene for downregulation.

1. Presented By:
Kunj Bihari Gupta
M.Phil-Ph.D (1st sem)
Center for Biosciences
School of Basic and Applied Sciences
Central University of Punjab

2. What are MicroRNAs (miRs)? –
 MicroRNAs are a class of 21-24 nucleotides long,
single stranded, endogenous, non-coding RNAs
involved in PTGS in all eukaryotes .
 Transcribed from genomic DNA mainly by RNA
poly II.
 Nascent transcript known as pri-miR, then premiR and finally form mature miR.
 Generate dsRNA with the target mRNA which
is chopped by different nucleases.

3.  First miR lin-4 was discovered in 1993 by Victor
Ambros in C. elegans
 Regulate lin-14 protein.
 In 2000 second miR (let-7) characterized in
C. elegans
 Total 24,521 miRs discovered so far.
MicroRNA Database. (2013). (http://www.mirbase.org/)

4.  According to standard nomenclature system, name of




any MicroRNA is written as mir-123.
miR = MicroRNA (mature form).
mir = Precursor MicroRNA.
Number indicates order of discovery.
Annotated with an additional lower case letter e.g.miR-123a & miR-123b, if deference in only one or two
nucleotides.

5.  Additional dash-number suffix e.g., miR-123-1 & miR-
123-2, if produce same miR but transcribed from
different places in the genome of same organism.
 When two miRs originate from opposite arms of the
same mir, then they are denoted with a -3p or -5p
suffix.
 When relative expression levels are known, then an
asterisk (*) are use to differentiate e.g.- miR-123 (more)
and miR-123*(less).

6. Species specific miR are denoted as below  hsa = Homo sapiens (Human)
 oar = Ovis aries (Sheep)
 ath = Arabidopsis thaliana (model plant)
 d = Drosophila

7. Genomic DNA
Transcription (Poly II)
In Side Nucleus
Pri-miRNA
Drosa+DGCR8 complex
Pre-miRNA
Exportin- 5
Pre-miRNA
In Side Cytoplasm
Dicer
Mature-miRNA

8. Nephew k.p. et al., 2010

9. Although the miRNA is only 24 nucleotides long,
But its 5' and 3' ends seem to have distinct roles• miRNAs can regulate their targets simply by strong
pairing with so-called seed sequence that consist of just
seven or eight bases complementary to the miRNA 5' end.
•miRNA 3' ends provide an additional measure of
regulatory control by permitting the function of target
sites that have only limited complementarity to the
miRNA 5' end.

10. Sun et al., 2010

11. miR can regulate gene expression in two ways:-
1. Indirect- activating/deactivating regulatory elements.
2. Direct- making double stranded transcript by binding
to regulating genes.

12.  miRNA is regulated by transcription factors (TFs) which
bind to few kb to more than 50 kb upstream of the miRNA
genes.
 Very few exact TF binding sites have been identified
experimentally.

13. • miRBase:
• Diana Lab:
http://www.mirbase.org/
http://diana.cslab.ece.ntua.gr/
•
•
•
•
•
•
•
•
http://www.ebi.ac.uk/enrightsrv/microcosm/
MicroCosm:
miRNAminer:
miRviewer:
Patrocles:
microRBase:
PicTar:
TargetRank:
TargetScanS:
http://groups.csail.mit.edu/pag/mirnaminer
http://people.csail.mit.edu/akiezun/miRviewer
http://www.patrocles.org/
http://microrna.sanger.ac.uk/
http://pictar.mdc-berlin.de/
http://hollywood.mit.edu/targetrank
http://www.targetscan.org/

14.  When plants are subjected to other than ideal growing


-
-
conditions, then it considered to be under stress.
Plants are sessile hence cannot escape from stress.
Stress may trigger a wide range of plant responses:Altered gene expression
Cellular metabolism
Changes in growth rates and crop yield, etc.

16.  Biotic – Imposed by another living organism
 Abiotic – Arising from an excess or deficit in the
physical and chemical environment.
BIOTIC STRESSES
Caused by living
organisms
•Fungi
•Bacteria
•Insects
•Herbivores
•Other plants/competition
ABIOTIC STRESSES
Environmental, nonbiological
•Temperature (high / low)
•Water (high / low)
•Salt
•Radiation
•Chemical
•Nutrition deficiency

17.  Function:
Stimulates early growth and
formation, hastens maturity, promotes
production and makes plants hardy.
root
seed
 Symptoms: Small root growth, spindly stalk, delayed
maturity, purplish discoloration of leaves, dying of tips
of older leaves, and poor fruit and seed development.

18. Nilssona L et al., 2010

19.  Ubiquitin-conjugating E2 enzyme (UBC24).
 Phosphate starvation responsive (PHR1, PHR2).
 Phosphate transporter genes (Pht1.8 & Pht1.9).
 PHO1,PHO2 gene. etc.

20. Nilssona L et al., 2010

22. Root
Shoot
Hsieh Li-Ching et al., 2009

23.  Out of many miR, miR399 are most studied.
 Which is strongly induced upon P starvation.
 It is a family of nearly similar 6 different miR named as
miR399a,
miR399b
miR399c,
miR399d,
miR399f in case of Arabidopsis.
 The target for this miR is PHO2 gene.
 Binds to the 5’ UTR of PHO2 transcripts.
miR399e,

24. Conclusion
 Plants are sessile in nature so, continuously exposed to
different stress.
 It is interesting to know the role of miRNAs, involved
in the regulation of plant abiotic stress.
 To find out stress-responsive miRNAs, smallRNA
libraries were created and sequenced with pooled
RNAs from plants treated with different abiotic
stresses

25.  miR399 are most studied in P starvation state.
 This miR down regulate the PHO2 gene transcripts.
 Expression of this miR is also tightly regulated by an
transcription factor MYB-2 in Arabidopsis.
Sun et al., 2010

26. Baek, D., Park, H. C., Kim, M. C.& Yun, D.-J. (2013). The role of Arabidopsis MYB2
in miR399f-mediated phosphate-starvation response. Plant Signaling & Behavior
8(3): 234-238.
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