Cloning and characterization of drought stress-induced NAC transcription factors from Brassica juncea and Sinapis alba

Abstract

The plant specific NAC [for NAM (no apical meristem),
ATAF, CUC (cup-shaped cotyledon)] TFs are one of the
largest plant TF families that play important roles in plant
development and stress tolerance. Suppression subtractive
hybridization (SSH) analysis with using drought stressed
plants of S. alba lead to the identification of several stress
responsive ESTs. Two of them homologous to Arabidopsis
NAC14 and NAC19 were selected for cloning of full length
CDS and expression analysis in Brassicaand related species
with contrasting drought tolerance. NAC14 and NAC19
genes were cloned from drought tolerant Sinapis alba and
Brassica juncea cvs. RGN73 and Varuna, and drought
sensitive B. juncea cvs. RLM619, BEC144 and BioYSR.
Sequencing of genomic region coding for these NACs
revealed that both NAC14 and NAC19 contain 3 exons and
2 introns each. In silico analysis of protein structure led to
development of 3D models of these stress responsive NAC
TFs. Although both proteins have 7 strands of β sheets,
the NAC14 had 5 β sheets of type A and 2 β sheets of type
B, while NAC14 have all 7 β sheets of type A. These proteins
also differed in helix content, β turns and g turns. This
suggest their functional diversity under abiotic stresses.
Real-time RT-PCR expression analysis revealed that both
the genes were up-regulated under drought stress in the
leaves of B. juncea genotypes Varuna and BioYSR. In
addition, NAC14 was up-regulated in the leaves of RLM619,
while NAC19 was up-regulated in the leaves of S. alba and
BEC144 under drought stress as compared to control
conditions. Interestingly, drought stress did not up-regulate
these genes in RGN73. This study revealed genotypic
variation in the drought regulation of NAC TFs in B. juncea
and S. alba.