different plant species and RNA-Seq, Notaguchi et al. [8] identified
138 transcripts of Arabidopsis that were mobile across an Arabidopsis/tobacco graft union. Approximately ten of these were
RNAs that encode transcription factors (TFs), including transcripts
from a BEL1-like and a KNOX-type TFs. In another study, using
variant ecotype transcript profiling in heterografts, 2006 genes
producing mobile RNAs were identified in Arabidopsis [9]. Many
of these mobile transcripts followed the phloem-dependent sugar
allocation pathway from leaves to roots, but a high number of
transcripts also moved in a root-to-shoot direction. Despite these
valuable insights, mobile RNAs with an established function or
phenotype are still rare. Some of the best examples of these latter
types include StBEL5, À11, À29 [10, 11], and POTH1 [12] of
potato; CmGAI of pumpkin [13]; PFP-LeT6 from tomato [14];
and AUX/IAA [15], FLOWERING LOCUS T (FT), and CENTRORADIALIS [16–18] from Arabidopsis. Molecular tools for
assaying movement are critical for elucidating the function of these
numerous phloem-mobile transcripts.
Potato virus X (PVX), a member of the Potexvirus genus, has a
positive-sense single-stranded RNA genome encoding five open
reading frames (ORFs) [19, 20]. The first ORF at the 5
0 -terminus
encodes the 166 kDa RNA-dependent RNA polymerase gene and
the last one at the 3
0 -terminus for the coat protein (CP). Between
them is the triple-gene block of three overlapping ORFs, which
encode proteins of 25, 12, and 8 kDa (Fig. 1). These three proteins
and the CP are required for cell-to-cell movement, and CP is
important for long-distance spread of PVX; but these proteins are
dispensable for replication. The p25 protein is also involved in
suppression of the antiviral RNA silencing defense mechanism
[21]. PVX has been modified as a plasmid vector that is widely
used for foreign gene expression and as a functional tool to activate
RNA silencing that specifically targets and suppresses foreign RNA
invading plant cells [22, 23]. It has also been used to monitor RNA
movement [24].
Two mobile RNAs that have been clearly implicated in regulating developmental processes are FT in floral induction and StBEL5
in tuber formation. The regulatory RNA movement sequences
have been identified in both. FT movement depends on a stretch
of 102 nucleotides (nt) present in the coding sequence [24] and for
StBEL5, the mobility sequence consists of specific cytosine/uracil
(CU) motifs present in the 3
0 untranslated region (UTR) [10, 25,
26]. In potato, specific RNA-binding proteins, designated StPTB1
and StPTB6, bind to the CU motifs present in the 3
0 UTR to
stabilize and mobilize transcripts of StBEL5 [26]. Based on this
information, an RNA mobility assay was developed using the PVX
vector to investigate long-distance signaling by FT mRNA [16, 24]
and StBEL5 mRNA [10] movement in flowering induction and
tuberization, respectively. In both cases, deletion of the coat
182
Zhiming Yu et al.
138 transcripts of Arabidopsis that were mobile across an Arabidopsis/tobacco graft union. Approximately ten of these were
RNAs that encode transcription factors (TFs), including transcripts
from a BEL1-like and a KNOX-type TFs. In another study, using
variant ecotype transcript profiling in heterografts, 2006 genes
producing mobile RNAs were identified in Arabidopsis [9]. Many
of these mobile transcripts followed the phloem-dependent sugar
allocation pathway from leaves to roots, but a high number of
transcripts also moved in a root-to-shoot direction. Despite these
valuable insights, mobile RNAs with an established function or
phenotype are still rare. Some of the best examples of these latter
types include StBEL5, À11, À29 [10, 11], and POTH1 [12] of
potato; CmGAI of pumpkin [13]; PFP-LeT6 from tomato [14];
and AUX/IAA [15], FLOWERING LOCUS T (FT), and CENTRORADIALIS [16–18] from Arabidopsis. Molecular tools for
assaying movement are critical for elucidating the function of these
numerous phloem-mobile transcripts.
Potato virus X (PVX), a member of the Potexvirus genus, has a
positive-sense single-stranded RNA genome encoding five open
reading frames (ORFs) [19, 20]. The first ORF at the 5
0 -terminus
encodes the 166 kDa RNA-dependent RNA polymerase gene and
the last one at the 3
0 -terminus for the coat protein (CP). Between
them is the triple-gene block of three overlapping ORFs, which
encode proteins of 25, 12, and 8 kDa (Fig. 1). These three proteins
and the CP are required for cell-to-cell movement, and CP is
important for long-distance spread of PVX; but these proteins are
dispensable for replication. The p25 protein is also involved in
suppression of the antiviral RNA silencing defense mechanism
[21]. PVX has been modified as a plasmid vector that is widely
used for foreign gene expression and as a functional tool to activate
RNA silencing that specifically targets and suppresses foreign RNA
invading plant cells [22, 23]. It has also been used to monitor RNA
movement [24].
Two mobile RNAs that have been clearly implicated in regulating developmental processes are FT in floral induction and StBEL5
in tuber formation. The regulatory RNA movement sequences
have been identified in both. FT movement depends on a stretch
of 102 nucleotides (nt) present in the coding sequence [24] and for
StBEL5, the mobility sequence consists of specific cytosine/uracil
(CU) motifs present in the 3
0 untranslated region (UTR) [10, 25,
26]. In potato, specific RNA-binding proteins, designated StPTB1
and StPTB6, bind to the CU motifs present in the 3
0 UTR to
stabilize and mobilize transcripts of StBEL5 [26]. Based on this
information, an RNA mobility assay was developed using the PVX
vector to investigate long-distance signaling by FT mRNA [16, 24]
and StBEL5 mRNA [10] movement in flowering induction and
tuberization, respectively. In both cases, deletion of the coat
182
Zhiming Yu et al.