Expression of ATFL1 correlates with the induction of
flowering in C. pallens
Leaf samples from the control plants at 1070 m and transplants that
flowered in the next season collected during and after the
transplantation period were analysed for the expression of selected
flowering-time genes (Table 1).
Leaf samples collected from both the 17Hot transplants that flowered in
2018 and control plants (17control) that had remained vegetative showed
a seasonal expression pattern (P < 0.05; Table S5; Fig. S2).
The seasonal expression patterns for CpFT3, CpFT4, CpFT5, CpATFL1,
CpVRN1, CpMADS50, CpMADS1 and CpHd1 (Fig. S2) were similar
between the control plants at 1070 m, all of which had remained
vegetative, and the transplants that flowered in the next season (Table
S5). Greater expression of FT-like genes was observed in the
spring season (October samples) aligning with studies in model plant
species (Nagano et al., 2019). Expression of FT-like genes was
similar during the spring season in both leaf sample sets, whether from
tillers that flowered in the next season or remained vegetative.
However, CpFT1 transcript was not detected in any of the samples
due to its expression either being below the limit of detection or
because it was not expressed in the leaves of the plant during the time
when leaf samples were collected. Expression analysis of CpVRN1(Fig. S2), another floral promoter, correlated with its known seasonal
expression pattern in temperate cereal species (Woods, Ream, & Amasino,
2014).
When the expression pattern is compared between the two sets of plants
during the inductive summer conditions (January 2017), CpATFL1had a significantly greater expression in the leaf samples of the 17Hot
transplants that flowered in the next season compared to the control
plants at 1070 m that remained vegetative. Along with CpATFL1,other floral promoters including CpTPS1, CpMADS1, CpMADS50, andCpVRN1 were also highly expressed in the leaf samples associated
with flowering tillers (Fig. 2a, Fig S3).
The expression of flowering-time genes in the leaf samples from the
transplanted plants that had flowered in 2017 (16Hot transplants and
16Cool transplants) was also compared to the control plants at 1070 m
(16Control), none of which flowered. All the gene(s) showed a seasonally
variable expression pattern (P < 0.001; Table S5) (Fig. S4).
The expression pattern of CpFT2 , CpFT3 , CpFT4 ,CpATFL1 , CpMADS1 , CpEhd3 and CpTPS1 in the
leaves of tillers that flowered in the next season at both the sites, UC
and 1520 m (16Hot transplants and 16Cool transplants, respectively)
showed a similar expression pattern to that observed in the leaf samples
associated with flowering in 17Hot transplants (comparing Fig. S3 and
Fig. S4). Even though CpFT5 had a seasonal expression pattern,
plants at UC showed greater expression during autumn. All the genes
showed a significant differential expression between tillers which
subsequently flowered vs tillers that remained vegetative during the
induction summer period (P < 0.001), except for CpFT4in the 16Cool transplants (Table S5).
Similar to the 17Hot transplants, CpATFL1 had a significantly
greater expression in the tillers that flowered at both the sites (UC
and 1520 m; 16Hot transplants and 16Cool transplants, respectively)
during the inductive summer period (January-2016) (Fig. 2a). Expression
analysis of CpVRN1 also correlated with its known seasonal
expression pattern with a higher peak in expression post winter (Shimada
et al., 2009). The expression of CpVRN1 was greater in the
tillers at both the sites (UC and 1520 m) that flowered in the next
season compared to the plants at the control site that had remained
vegetative (Fig. S3). Expression of CpHd1 , CpGI ,CpEhd3 , CpMADS50 and CpMADS1 were all greater in
the leaf samples of the transplanted plants which subsequently flowered
compared to the control site plants that remained vegetative (Fig. S3).
Leaf samples associated with tillers that remained vegetative in both
the transplants, (16Hot transplant and 16Cool transplants) were found to
have significantly lower expression of CpATFL1 during the
inductive summer temperature (January 2016) compared to the leaf samples
from the same groups of transplants that flowered in the next season
(Fig. 2b). In addition to the transplants, leaf samples collected from
the control plants during January 2018 (18 Control) that flowered in the
masting year 2019, also showed a significantly greater expression ofCpATFL1 compared to the leaf samples from plants that remained
vegetative in the year 2019 (Fig. 2b). In summary, expression studies
suggest that an elevated expression of CpATFL1 during the
inductive summer temperature period is associated with the induction of
flowering in C. pallens .