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Biology and Biotechnology of Environmental Stress Tolerance in Plants, Volume 3

9.5 BIOGENESIS OF SMALL RNA

The biogenesis of various sRNAs in plants, such as miRNAs and siRNAs,

is quite similar. Both of these sRNAs are made from larger RNA precursors

by enzymes termed DICER-LIKE, which are ribonuclease-III-like nucleases

(DCL). In the context of sRNA biogenesis in animals, the nuclease is known

as Dicer. However, there are some basic differences in their processing.

Usually, miRNA is produced from the genetic loci called MIR genes (Figure

9.1). Just like other protein-coding genes MIR genes are also first transcribed

into a primary transcript (pri-miRNA), which then forms an incomplete

“fold-back” stem-loop or hairpin structure due to the existence of intra­

molecular sequence complementarity (Guo et al., 2016). The transcription

is generally mediated by RNA polymerase II (RNA Pol-II) to yield primary

miRNA (pri-miRNA) which is subsequently subjected to 5’ capping and 3’

polyadenylation (Koroban et al., 2016). In the nucleus, from this pri-miRNA

structure, DCL1 produces a pre-miRNA molecule with a specific “stem­

loop” structure in collaboration with the dsRNA-binding protein DRB1 or

HYPONASTIC LEAVES 1 (HYL1) (Eamens et al., 2009; Kurihara et al.,

2006; Kurihara & Watanabe, 2004). According to Voinnet (2009) DCL1,

HYL1 and SERRATE proteins have collaboratively executed the processing

of pre-miRNAfrom the primary transcript (pri-miRNA) in plants. The further

process of pre-miRNA is mediated by DCL1 within the nucleus, generating

a 21 nucleotide long imperfect RNA duplex that is comprised of mature

miRNA (guide strand) and newly synthesized miRNA (miRNA rider stand).

The RNA methylase HUA ENHANCER1 (HEN1) then methylates the 3’

terminal nucleotides of the RNA duplex at the 2’-O-hydroxyl group, which

facilitate to shield of the miRNA: miRNA duplex from degradation (Li et

al., 2005). After that, an exportin protein HASTY (HST1) release the duplex

into the cytoplasm. Inside the cytoplasm, the strand of miRNA duplex which

is integrated into the AGO protein regulates enzymatic activity of RISC and

directs RISC to connect with related target transcripts through sequence

complementarity (Achkar et al., 2016; Kim et al., 2009; Lee et al., 2004).

The siRNA molecules are generated from dsRNAs which may be endog­

enous or exogenous in origin. Plants generate various kinds of siRNAs viz.,

trans-acting siRNAs, phased secondary siRNA, natural antisense siRNA,

and hc-siRNA. Endogenous sources of siRNAs include dsRNA generated

through natural cis-antisense, dsRNA produced from heterochromatin, gene

paired encoded mRNAs and also as an outcome of miRNA induced cleavage

of ssRNAs (Chinnusamy et al., 2007). Among the exogenous origins of