A forward genetic approach to isolate <em>Arabidopsis</em> mutants altered in the regulation of the <em>ALDH7B4</em> gene

Abstract

The ALDH7B4 gene is a member of the ALDH super gene family which is known to be involved in stress tolerance, but the regulation of ALDHs was unknown. In the current studies, a forward genetic approach was used to propose potential regulators for ALDH7B4. For this, a homozygous transgenic <em>Arabidopsis</em> line carrying a single copy of the ALDH7B4promoter-GUS construct was mutagenized with EMS. The mutant population was screened for mutants with aberrant GUS expression. Two mutants F38 and B10 were selected on the basis of high and low GUS expression, respectively. The mutants were characterized to assess the effect of the mutations on the physiological, molecular and genetic levels. The mutants showed contrasting phenotypes from each other for most of the features studies. The F38 mutant is hypersensitive to NaCl, mannitol, glucose and ABA for radicle emergence and cotyledon formation while B10 showed reduced sensitivity to these stressors. A correlation between differential expression of ALDH7B4 and dormancy has been found in the mutants (higher in the F38 and lower in B10). This points to a role of ALDH7B4 in seed dormancy and germination. ABA is a principal factor on the basis of expression of NCED6, a rate limiting enzyme of the ABA biosynthesis pathway, and ABA signaling genes (ABI3 and ABI5) for inducing primary seed dormancy. The regulation of ALDH7B4 seems to be controlled in an ABA-dependent fashion both under salt and drought stress. Lower accumulation of malondialdehyde (MDA) and reactive oxygen species (ROS) under salt and drought stress suggests that the B10 mutant is a stress tolerant line. Stable contents of chlorophyll under salt stress in the F38 mutant implies that the photosynthetic apparatus of the F38 mutant is salt tolerant. Genetic analysis unveiled that the B10 mutation(s) is dominant for decreased ABA sensitivity and controlled by nuclear gene(s). The EMS mutations reduced the root length of the F38 mutant while the activity of the ALDH7B4 promoter was increased in theses roots which suggest a role of ALDHs in root development. Genetic analysis revealed that the F38 mutations are recessive and heterosis (94.25%) controlled by nuclear genes is involved. I propose four most potential regulators of ALDH7B4 on the basis Whole Genome Sequencing of the F38 short rooted mutant plants.