## 3 Discussion
### TrHSFB2a Negatively Regulates Drought, Heat and Salt Stress in Arabidopsis  
The overexpression of _TrHSFB2a_ in _Arabidopsis_, although it enhanced the transcriptome abundance and RNA interference of _AtHSFB2a_ in _Arabidopsis_, reduced the accumulation of _AtHSFB2a_ transcripts. There were no significant differences in plant stature, root length or fresh and dry weights under normal conditions. It indicated that _TrHSFB2a_ does not have significant effects on plant development under normal conditions, as similarly observed in the effects of HSBs of class B on other plant species [40; 41]. However, under drought, salt and heat stress, the overexpression of _TrHSFB2a_ significantly reduced plant growth compared to wild type, while RNA interference of _AtHSFB2a_ significantly improved salt, heat and drought tolerance both in 1/2 MS media and in the soil. By increasing the stress levels (NaCl 150 mM and Mannitol 300 mM) or by extending stress duration, the overexpression of _TrHSFB2a_ led to significantly higher drying and dying rates compared to wild type, while RNA interference of _AHHSFB2a_ exhibited opposite trends. These observations agreeing has been observed in the transgenic lines developed by overexpression and RNA interference homolog of this _OsHSFB2b_ in rice [41] and overexpression of _CarHSFB2b_ of Chickpea in Arabidopsis [40]. RWC, photosynthetic activity, REC percentage and MDA contents are important indicators to assess the stress tolerance potential differences in plants. Under drought and heat stress treatment, the overexpression lines of _TrHSFB2a_ showed higher REC % and MDA contents than that of wild type, and in T-DNA lines of _AHHSFB2a_, the accumulation of MDA contents and REC % was lower compared to wild type. As electrolyte leakage is an indirect indicator of cell membrane damage due to stress conditions [45; 46], the higher REC in _TrHSFB2a_ overexpression lines showed higher cell membrane damage, which had occurred there, while there was comparatively less cell membrane damage in T-DNA lines. One of the end products of lipid peroxidation due to external stresses is MDA, which is a result of membrane damage caused by free radicals [47; 48], was higher in overexpression lines and lower in RNA interference lines compared to wild types, representing higher lipid damage in overexpression lines and a lower in T-DNA lines. Abiotic stresses on plants mainly reduce the photosynthetic efficiency of the plants under stress as a result of the negative effects of the abiotic stresses on the biosynthesis of photosynthetic pigments, photosystem performance, CO2 fixation and gases exchanges, and on carbohydrate metabolism, etc. The photosynthetic activity and photosystem performance index significantly reduced overexpression lines and significantly improved in T-DNA lines compared to wild-type user drought, salt and heat stress, indicating better adaptability of T-DNA line and reduced fitness by overexpressing _TrHSFB2a_ gene in _Arabidopsis_. Moreover, lower RWC, as observed in overexpression lines, can reduce or even completely stop photosynthetic activity in a stressed condition. Leaf wilting, drying, RWC, photosynthetic system performance, REC and MDA-related to plant stress that has been already used to assess the function of HSFs under drought, salt and heat stress in transgenic plants [41; 49; 50]. These findings provide evidence that _TrHSFB2a_ negatively regulates stress tolerance at least in _Arabidopsis_. The HSFB class has been reported to act as a transcriptional repressor in plant species [23; 26; 41], including _Arabidopsis_[5; 26]. In _Arabidopsis_, the single mutant _HSFB2B_ and double mutant _HSFB1 HSFB2_ induced the strong expression of stress defense-related genes, and thereby, significantly enhanced the resistance of _Arabidopsis_ against pathogens [27]. These double mutants also improve heat tolerance in _Arabidopsis_, which might be because _HSFB1_ and _HSFB2b_ repress the expression of heat shock (HS) inducible genes required in the stress tolerance process [26].