4.4 Molecular studies
The results of gene expression analysis showed that allelochemical compounds altered the expression of PsbA and PsbS genes directly or indirectly. Interestingly, these changes were dependent on plant species; so it seems that the strategy and ability of each plant for tolerating allelopathic stress determines the fate of changes in gene expression. Increasing PsbA gene expression in wheat plant (which encodes D1 subunit of photosystem II due to increasing chlorophyll fluorescence) declines the use of reduced coenzymes, reduction of stomatal opening, and subsequently reduction of CO2input and carbon fixation and ultimately reduction of photosystem II efficiency and increasing risk of D1 subunit degradation in this photosystem are quite obvious. In fact, by continuously replacing degraded D1 protein, wheat plant retains the production of NADPH/H+ and ATP to fixate respiratory carbon and avoid water loss, whereas the decline in cucumber’s PsbA gene expression caused cucumber not to be able to continuously replace degraded D1 protein. According to the increasing chlorophyll fluorescence in this plant and extreme risk of D1 subunit degradation, cucumber plant increased the stomatal opening and photosynthesis rate to reduce the excess electron in the photosynthetic electron transport chain. The results of PsbS gene expression confirm the interpretations provided for both plants. The expression of this gene was decreased in both plants, but it was statically significant (p ≥ 0.01) only in cucumber plant (99.93%). Decline in PsbS gene expression led to the destruction of the optical antenna components. Destruction of antenna structure, reducing the energy supply for excitation of reaction centers, especially in photosystem II, led to better control of excess electron entry into the photosynthetic electron transfer chain. The results of molecular docking showed that the proteins encoded by PsbA and PsbS can interact directly with thymol, carvacrol, beta-elemene, trans-caryophyllene, germacrene-D, spathulenol, and 5alpha-pregnane compounds. These compounds are well-known allelochemicals in plants. Therefore, it can be concluded that allelochemicals not only indirectly (such as induction of oxidative stress) but also directly are able to decrease protein’s function by binding to their structure. All of the above mentioned compounds have the ability to bind to proteins encoded by the PsbA and PsbS genes, and among them the 5alpha-pregnane found in wheat shoot has the best structure for this binding.