Despite previous studies on exploring the environmental effects of titanium dioxide nanoparticles particle (nTiO2) on plants, the detailed impacts of nTiO2 on the antioxidant system and photosynthesis of plants is still not well understood. This study was aimed at investigating the physiological and biochemical responses to nTiO2 by oxidative damage, Ti bioaccumulation, cell death, and photosynthesis in wheat. The results showed that 5.0 g nTiO2 L−1 resulted in a significant decrease in plant growth, chlorophyll contents, and photosynthetic activity. However, the obvious accumulation of reactive oxygen species (ROS) and cell death were observed under nTiO2 treatments in wheat roots and leaves. In addition, the concentrations of Ti in the roots were significantly higher than that in leaves with increased nTiO2 concentrations. Significant increase in enzyme activities and the levels of ascorbate were found in leaves exposed to 1.0 and 5.0 g nTiO2 L−1. Furthermore, the level of D1 and PsbS remarkably decreased in wheat leaves at 5.0 g nTiO2 L−1. However, the strong phosphorylation of photosystem II (PSII) reaction center protein D1 and D2 was observed at 5.0 g nTiO2 L−1. Altogether, these findings demonstrated that the roots suffered from more severe toxic damage from nTiO2 than the leaves and wheat plants respond to nTiO2 through the different physiological and biochemical mechanisms in the roots and leaves.