Understanding molecular response of Arabidopsis to salt, heat and highlight stress combinations.

Plants in nature are exposed to combinations of abiotic and biotic stresses. Climate change driven increase in frequency of abiotic stress combinations is being detrimental to the plant survival and growth. Previously we found that Arabidopsis plants when exposed to combination of abiotic stresses termed as multifactorial stress combination (MFSC), the accumulation of multiple mild abiotic stresses becomes damaging to plants causing severe reduction in plant survival which otherwise not detrimental when exposed as single stresses. Here in this study, we investigated the role of a transcription factors Basic helix loop helix (bHLH35) in MFSC tolerance in Arabidopsis seedlings. In this study, Arabidopsis thaliana wild-type (Col0), and mutants of bHLH35 gene plants were subjected to salt stress (S; 50mM), high light stress (HL; 700 µE), heat stress (HS; 33 °C) and different combinations of these three stresses (S+HL, S+HS, HL+HS, S+HL+HS). Sterilized seeds of above-mentioned genotypes were germinated on ½ MS (murashige and Skoog media) with or without salt, at 21°C temperature and 150 µE light intensity. 7 days old seedling were subjected to HL and HS for 3 days (in combinations mentioned above) to determine the impact of stresses on survival. Seedlings for control were grown on ½ MS media without salt at 21°C temperature and 150 µE light intensity. To understand the transcriptomic response of Arabidopsis genotypes, 7 days old seedling growing on ½ MS with or without salt were exposed to HS and HL stress (in combinations mentioned above) for 1.5 hrs and seedlings from all the stress treatments and control were snap frozen in liquid nitrogen. Total RNA from the seedlings were isolated using Qiagen RNeasy® Plant Mini Kit. The RNAseq analysis was performed by Novogene Co. RNA libraries were prepared using standard Illumina protocols and RNA-Seq was performed using a NovaSeq 6000 PE150 platform by Novogene Co. Ltd. (https://en.novogene.com/; Sacramento, CA, USA). Read quality control was performed using Trim Galore v0.6.4 and FastQC v0.11.9.