Identification of the seed-specific MtHSFA9 in M.
truncatula as the ortholog of HaHSFA9
To identify transcription factors (TF) that underly the regulation of
the acquisition of desiccation tolerance (DT) and longevity, we
performed a detailed temporal transcriptome study during seed
development (Verdier et al. 2013). The construction of a gene
co-expression network of genes and seed specific transcription factors
that correlated with DT and longevity inferred a number of TFs as
putative regulators of late maturation/longevity genes (Verdier et
al. 2013). Detailed characterization of this network identified a heat
shock transcription factor (Medtr4g126070/HSFA) as a central hub of the
module, with the highest betweenness centrality (Fig. 1a, Data S1). In
addition, this HSFA was one of the nodes with the closest centrality and
number of connected nodes, with only Medtr1g023170, a DREB2D TF, being
more connected (Data S1).
A blast of the M. truncatula sequence against Arabidopsis TAIR11
revealed homology with three AtHSFA: At2g26150/HSFA2 (7.55e-75),
At3g22830/HSFA6B (1.25e-72) and AT5G54070/AtHSFA9 (7.17e-72). To analyze
to which group the M. truncatula HSFA belongs, a phylogenic tree
was constructed using homologous HSFA protein sequences retrieved fromM. truncatula, Glycine max and Lotus japonica databases
(Fig. 1b). The phylogenetic reconstruction was performed using maximum
parsimony and identified Medtr4g126070 as part of a subgroup containing
AtHSFA9, HaHSFA9, GmHSFA9.1 and GmHSFA9.2 (Fig. 1b). We therefore
referred to the gene as MtHSFA9 . Since the Arabidopsis and
sunflower HSFA9 are specifically expressed in seeds, we investigated the
expression patterns of MtHSA9 and the other HSFA genes
(Fig. 1c). MtHSFA9 is predominantly expressed in seed tissues,
with the highest transcript levels at the end of seed maturation (36
days after pollination (DAP)) (Fig. 1c, Fig. S1a). A moderate level of
transcript was also detected in shoots submitted to severe drought.