Objectives This study aimed to elucidate the mechanism by which phenylalanine ammonia-lyase inhibitor (AIP) modulates root endodermal differentiation and cadmium (Cd) accumulation in rice, providing a theoretical basis for breeding low-Cd-accumulating rice cultivars.

Methods Hydroponic experiments were conducted using a low-Cd japonica rice cultivar Nipponbare and a high-Cd indica rice cultivar IR32307. Initially, we investigated the effects of AIP (5 and 10 mmol/L) on root phenylalanine ammonia-lyase (PAL) activity under both Cd stress and non-stress conditions to determine the optimal AIP concentration for subsequent experiments. Subsequently, hydroponic experiments were conducted with four treatments: a control (CK) without AIP or Cd stress, a 10 mmol/L Cd stress treatment (Cd), a 10 mmol/L AIP treatment (AIP), and a combined Cd+AIP treatment. After treating the rice seedlings for one week, Cd content in the shoots and roots was determined using inductively coupled plasma mass spectrometry (ICP-MS). The development of the endodermal barrier was observed via laser confocal microscopy, and Cd transport pathways were analyzed using short-distance transport tracing techniques. Additionally, quantitative real-time PCR (qPCR) was employed to measure the expression levels of genes related to endodermal differentiation.

Results Cd stress significantly induced PAL activity in rice roots, with increases of 107.49% and 21.45% in Nipponbare and IR32307, respectively, relative to the CK. This induction promoted lignin and suberin deposition and upregulated the expression of OsPAL, OsCASP1, OsCYP86A1, and OsKCS20, thereby enhancing endodermal barrier development. Conversely, following the inhibition of PAL activity by AIP (Cd+AIP treatment), lignin and suberin contents in the roots of both cultivars decreased by an average of 60.40% and 37.42%, respectively, compared with the Cd treatment alone. Relative to CK, AIP treatment delayed the development of the casparian strip and suberin lamellae, as evidenced by significant increases in root D_TIP-CSs (16.62% in Nipponbare; 16.41% in IR32307) and D_TIP-SL (14.10% in Nipponbare; 8.95% in IR32307), resulting in weakened endodermal barrier function. Concurrently, AIP treatment increased Cd accumulation in the shoots and roots of Nipponbare by 27.37% and 4.36%, respectively, with IR32307 exhibiting a similar trend.

Conclusions PAL-mediated lignin and suberin synthesis constitutes a core regulatory link in the development of the root endodermal barrier. Nipponbare exhibited a stronger capacity for PAL activity induction and earlier endodermal differentiation compared to IR32307; these physiological and structural characteristics serve as the key basis for its restriction of Cd uptake and subsequent translocation to the shoots.