Phosphorus (P) is an essential nutrient for plant growth. In recent decades, the application of phosphate (Pi) fertilizers has contributed to significant increases in crop yields all over the world. However, low efficiency of P utilization in crops leads to intensive application of Pi fertilizers, which consequently stimulates environmental pollution and exhaustion of P mineral resources. Therefore, in order to strengthen the sustainable development of agriculture, understandings of molecular mechanisms underlying P efficiency in plants are required to develop cultivars with high P utilization efficiency. Recently, a plant Pi-signaling network was established through forward and reverse genetic analysis, with the aid of the application of genomics, transcriptomics, proteomics, metabolomics, and ionomics. Among these, proteomics provides a powerful tool to investigate mechanisms underlying plant responses to Pi availability at the protein level. In this review, we summarize the recent progress of proteomic analysis in the identification of differential proteins that play roles in Pi acquisition, translocation, assimilation, and reutilization in plants. These findings could provide insights into molecular mechanisms underlying Pi acquisition and utilization efficiency, and offer new strategies in genetically engineering cultivars with high P utilization efficiency.