4.1 PpINHs are cold-responsive regulators that have evolved with
flowering plants
Plant invertase inhibiters are small proteins that have been identified
in a variety of plant species, such as tobacco (Greiner et al. 1998;
Weil, Krausgrill et al. 1994), Arabidopsis (Link et al.2004),
maize (Bate et al. 2004), tomato (Reca et al., 2008), potato (Brummell
et al., 2011), and sugarcane (Shivalingamurthy et al., 2018). Wan et al.
2018, have reported that INHs, have evolved with vascular plants; our
BLASTP results support this, as we did not identify any INHs from lower
plants such as algae and moss (Figure 1a, b ), and inAmborella trichopoda, the common ancestor of living angiosperms
(Albert et al., 2013), no INH gene was predicted, which supports
the thesis that INHs evolved in flowering plants. Although the number ofINH genes varies among species, INH gene duplication
appears to have followed species evolution, indicating their importance
in modern vascular plants. Amino acid sequence alignments of INHs from
different species showed that all contained four highly conserved
cysteine (Cys) residues, a hallmark of plant INHs (Rausch, & Greiner,
2004). The nucleic acid sequences however, vary among species, even
within a species (Figure 1c) .
CBF is a type of AP2/ERF transcription factor; it binds to CRT/DRE
cis- elements in the promoter region of target genes regulating
their expression. In wild-type Arabidopsis , 302 genes are induced
by low temperature, in Arabidopsis constitutively expressing CBF,
only 85 genes are induced by low temperature. Moreover, among genes
downstream from CBF, 8 were inhibited when CBF was overexpressed and
these 8 genes are down-regulated by low temperature (Vogel et al.,
2005). These results suggest CBF plays a key role in plant cold
response. We found numerous CRT/DRE cis-elements in 5 PpINHpromoters (Figure 2 ), indicating these PpINHs are
targets of CBF. Expression of PpINH1 was significantly reduced in
peaches during cold storage (Figure 6f ) indicatingPpINH1 is sensitive to cold stress. These data suggest that
low-temperature sensitive INHs may be regulated by CBF proteins.