Cobalt stress enhanced dendrobine-type total alkaloids biosynthesis of Trichoderma longibrachiatum UN32 through reactive oxygen species formation.

Abstract

Trichoderma longibrachiatum UN32 is a well-documented mutant strain known to produce dendrobine-type total alkaloids (DTTAs). It was serendipitously observed that the addition of Co2+ to the medium resulted in a notable enhancement in DTTAs production in the T. longibrachiatum UN32 strain, accompanied by an upregulating effect on the expression of antioxidase-related genes. Hence, the objective of the present work was to ascertain whether ROS (intracellular levels of hydrogen peroxide) induced by Co2+ treatment has a beneficial or detrimental impact on DTTAs biosynthesis. A comparison of the intracellular levels of hydrogen peroxide (H2O2) and DTTAs treated with CoCl2 and CH3COOH revealed that CoCl2 was the optimal inducer for investigating the relationship between ROS formation and DTTAs production. This was due to the observation that ROS formation was reduced by approximately 4% and DTTAs production was increased by 12.55% in comparison to the CH3COOH treatment. The physiological results revealed that the introduction of Co2+ resulted in the oxidative damage and activation of the expression of intracellular superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD). Furthermore, it was confirmed that ROS induced by Co2+ was beneficial to DTTAs production by adding exogenous ROS scavengers. The inclusion of all ROS scavengers, including vitamin C, tocopherol, melatonin, mannitol, and sesamol, resulted in a reduction in ROS accumulation and a concomitant decrease in DTTAs production. Specifically, the addition of melatonin at a concentration of 0.4mg/L demonstrated significant effects, resulting in a 32.53% (P < 0.01) decrease in ROS accumulation and a 45.22% (P < 0.01) reduction in DTTAs production. Subsequently, the timelines of accumulation of intracellular H2O2 and DTTAs content indicated that ROS are also crucial for normal fermentation without CoCl2 addition. Specifically, the proper H2O2 dose for DTTAs accumulation is between 8.82 and 18.86μmol/g. The present study offers the initial experimental evidence indicating that CoCl2 enhance DTTAs production during the culture of T. longibrachiatum UN32 via leading an increase in intracellular ROS, which is conductive to DTTAs production and can be inhibited by the ROS scavengers. Our results provide insights into the mechanistic study of DTTAs biosynthesis.