Biogenesis of Mammalian miRNA - MicroRNAs as Tools in Biopharmaceutical Production - page 22

which miRNA modulate protein abundance is through the repression of translation
by impairing translation initiation.
Existing evidence also suggests that miRNA function to repress translation at
a point following translational initiation. Using sucrose gradient fractionation to
examine polysomal distribution of mRNA, results indicate that both synthetic and
endogenous miRNA target messages may be associated with polysomes (Petersen
et al. 2006 ; Nottrott et al. 2006 ;Guetal. 2009 ). Furthermore, synthetic mRNA
containing a cap-independent IRES may be subject to repression by miRNA and
provides evidence that the regulatory step occurs at some point after the initiation
of translation (Petersen et al. 2006 ). In fact, numerous miRNA have been found
associated with polysomal fractions indicating that mature miRNA control protein
expression at a point beyond initiation and is consistent with a model wherein miRNA
may also bind to and repress target mRNA at a post-initiation step (Kim et al. 2004 ;
Nelson et al. 2004 ; Maroney et al. 2006 ; Kong et al. 2008 ). The precise mechanism
controlling the miRNA-induced repression via regulating elongation or termination
steps remains unknown.

2.8 Target Regulation by Altering Transcript Stability

Regulation of post-transcriptional gene expression by miRNA also likely occurs
through the modulation mRNA stability or turnover. Following recognition of the
target mRNA, miRNA promote the deadenylation and decapping of the message
resulting in its destabilization (Fabian et al. 2010 ). Deadenylation may be, at least in
part, dependent on the interaction between PABP and GW182 of the RISC thereby
impairing the interaction between PABP and eIF4G (Fabian et al. 2009 ). It does
not appear that deadenylation alone results in enhanced decay, but instead suggests
that deadenylation may be requirement for decapping and degradation by either a
3 -5 or 5 -3 exoribonuclease. Intriguingly, recent work using rabbit reticulocyte
lysate suggests that the poly(A) tail, but not the 5 -m 7 G cap, of target mRNA is
required for miRNA-mediated effects (Ricci et al. 2011 ). In fact, in these studies
using rabbit reticulocyte lysates, translational repression was independent of either
deadenylation or decreased mRNA stability. Despite examples of deadenylation
and decay occurring to varying degrees, the extent to which these processes play an
important role in mammalian cells remains unclear and may indicate that the primary
means of regulation, at least in mammals, is through a mechanism involved in the
regulation of translation initiation (Fabian et al. 2009 ).

2.9 Summary

Similar to protein-coding genes, miRNA gene expression is usually regulated through
the recruitment of RNA polymerase II to produce a primary miRNA transcript. This
nascent transcript undergoes nuclear processing to generate 60-80 nt pre-miRNA
transcripts that are then exported out of the nucleus and further processed by the