Cadmium-induced glycine-rich protein (cdiGRP) is a cell wall-associated factor that increases callose levels in plant vasculature. To better understand the cdiGRP/callose regulation system, we identified a tobacco protein, GrIP (cdiGRP-interacting protein, GrIP), that associates with cdiGRP and localizes at the plant cell wall. Constitutive overexpression of GrIP enhanced the accumulation of the cdiGRP protein and callose in vasculature-associated cells with or without treatment with cadmium ions. That GrIP gene expression was not affected by cadmium ions indicated that GrIP does not directly modulate the callose levels induced by the treatment. Instead, GrIP most likely functions by further elevating the accumulated amount of cdiGRP, the expression of which is up-regulated by the cadmium ions. Interestingly, the levels of cdiGRP mRNA were not affected by constitutive expression of GrIP, demonstrating that the enhancement in cdiGRP protein accumulation by GrIP overexpression occurs posttranslationally. Collectively, these observations suggest that GrIP interacts with cdiGRP and increases its level of accumulation; in turn, the elevated amounts of cdiGRP induce callose deposits in the plant cell walls. Therefore, GrIP and cdiGRP represent sequentially acting factors in a biochemical pathway that regulates callose accumulation in the plant vasculature.