问题描述:
英语翻译
The function of RNAi in the regulation of SPL3
Transcripts that contain a miRNA target site are frequently
silenced in an RDR6-dependent fashion when they are expressed
as transgenes,and it has been reported that miRNA-directed
cleavage sensitizes transcripts to transitive silencing (Parizotto et
al.,2004).Consistent with this report,we found that all of our
constructs with a functional miR156 target site were significantly
more susceptible to silencing than constructs that lacked this site.
This made it difficult to generate reporter lines and to maintain
these lines over many generations.It did not interfere with our
ability to identify reliable reporter lines,however,because
transgenes undergoing post-transcriptional silencing have a
different expression pattern than SPL3.Whereas the expression of
SPL3 increases with time,transgenes undergoing silencing are
either permanently silenced very early in shoot development,or
display progressively lower levels of expression during shoot
growth (de Carvalho et al.,1992; Glazov et al.,2003; Palauqui et
al.,1996; Vaucheret et al.,2004).Although it is clear that
transgenes expressing transcripts with miRNA-target sites are
often subject to RNAi,whether this is also true for endogenous
transcripts is less certain.miRNA-directed transitive silencing is
well documented in the case of transcripts that produce transacting
siRNAs (Allen et al.,2005; Yoshikawa et al.,2005).
However,most protein-coding transcripts that are cleaved by
miRNAs – including the miR156-regulatedtranscripts SPL9 and
SPL15 – are not affected by sgs3 or rdr6,and are therefore unlikely
to be targets of RNAi (Allen et al.,2005; Peragine et al.,2004).
The observation that SPL3 and SPL4 are elevated in these mutants
suggests that these genes either have features that make them
unusually susceptible to RNAi (e.g.the presence of the miR156
target site in their 3\2 UTR),or that they are regulated indirectly by
this mechanism – for example,by a transcription factor that itself
is a target of RNAi.We have no conclusive evidence for either
possibility.However,the observation that zip-1,rdr6-11 and sgs3-
11 have nearly identical effects on SPL3 expression (Fig.6A)
(Peragine et al.,2004) suggests that these genes regulate SPL3 by
the same mechanism.Because ZIP is not generally required for
RNAi (Hunter et al.,2003) or for the miR156-dependent
suppression of 35S::GUS-SPL3 (Fig.6C),this observation may
indicate that these genes regulate SPL3 expression indirectly
The function of RNAi in the regulation of SPL3
Transcripts that contain a miRNA target site are frequently
silenced in an RDR6-dependent fashion when they are expressed
as transgenes,and it has been reported that miRNA-directed
cleavage sensitizes transcripts to transitive silencing (Parizotto et
al.,2004).Consistent with this report,we found that all of our
constructs with a functional miR156 target site were significantly
more susceptible to silencing than constructs that lacked this site.
This made it difficult to generate reporter lines and to maintain
these lines over many generations.It did not interfere with our
ability to identify reliable reporter lines,however,because
transgenes undergoing post-transcriptional silencing have a
different expression pattern than SPL3.Whereas the expression of
SPL3 increases with time,transgenes undergoing silencing are
either permanently silenced very early in shoot development,or
display progressively lower levels of expression during shoot
growth (de Carvalho et al.,1992; Glazov et al.,2003; Palauqui et
al.,1996; Vaucheret et al.,2004).Although it is clear that
transgenes expressing transcripts with miRNA-target sites are
often subject to RNAi,whether this is also true for endogenous
transcripts is less certain.miRNA-directed transitive silencing is
well documented in the case of transcripts that produce transacting
siRNAs (Allen et al.,2005; Yoshikawa et al.,2005).
However,most protein-coding transcripts that are cleaved by
miRNAs – including the miR156-regulatedtranscripts SPL9 and
SPL15 – are not affected by sgs3 or rdr6,and are therefore unlikely
to be targets of RNAi (Allen et al.,2005; Peragine et al.,2004).
The observation that SPL3 and SPL4 are elevated in these mutants
suggests that these genes either have features that make them
unusually susceptible to RNAi (e.g.the presence of the miR156
target site in their 3\2 UTR),or that they are regulated indirectly by
this mechanism – for example,by a transcription factor that itself
is a target of RNAi.We have no conclusive evidence for either
possibility.However,the observation that zip-1,rdr6-11 and sgs3-
11 have nearly identical effects on SPL3 expression (Fig.6A)
(Peragine et al.,2004) suggests that these genes regulate SPL3 by
the same mechanism.Because ZIP is not generally required for
RNAi (Hunter et al.,2003) or for the miR156-dependent
suppression of 35S::GUS-SPL3 (Fig.6C),this observation may
indicate that these genes regulate SPL3 expression indirectly
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