A unique family of proteins associated with internalized membranes in protein storage vacuoles of the Brassicaceae

The protein storage vacuole (PSV) is a specialized organelle in plant seeds that accumulates storage proteins and phytate during seed development. In many plant species, such as tomato and tobacco, the PSV contains two types of microscopically visible intra-organellar inclusions: a large crystalline lattice of membranes and proteins, the crystalloid, and one or a few large phytate crystals, the globoids. In seeds of the family Brassicaceae, the PSVs lack visible crystalloids and have many small globoids dispersed throughout. We biochemically fractionated PSVs from Brassica napus and defined a crystalloid-like fraction that contained integral membrane protein markers found in crystalloids of other plants. Protein analyses identified a previously undescribed family of proteins, the Brassicaceae PSV-embedded proteins (BPEPs), associated with 'crystalloid' and globoid fractions. The defining characteristics of the BPEPs are an N-terminal signal peptide and tandem MATH domains, which may mediate protein-protein interactions. Database analyses indicated that the BPEPs are unique to Brassicaceae. Immunofluorescence studies using anti-BPEP antibodies and antibodies to other biochemical markers to label B. napus and Arabidopsis thaliana seed sections localized the BPEPs to structures within the PSVs, whose appearance was consistent with a diffuse network of internalized membranes and globoids. These results demonstrate that Brassicaceae PSVs contain internalized membranes, and raise the possibility that BPEPs modify these internal membrane structures to yield a PSV morphology different from that of tomato or tobacco.