Localization of repetitive proline-rich proteins in the extracellular matrix of pea root nodules

Summary Early responses of legume roots toRhizobium inoculation include new cell wall synthesis and induction of some putative wall protein genes. Although the predicted amino acid sequences of several early nodulins indicate that they encode proline-rich proteins (PRPs), the proteins have been neither isolated nor has their presence been demonstrated in cell walls. We have used polyclonal antibodies against PRP2 from soybean to identify and localize proline-rich proteins in pea nodules. On immunoblots, several PRPs were detected, ranging from less than 20 kDa to 110 kDa. Immunocytochemistry revealed that tissues of the vascular cylinder contained abundant PRPs, particularly in the secondary cell walls of xylem elements and phloem fibers. PRPs were also found within the primary wall of the nodule endodermis and within Casparian strips of the vascular endodermis. Of symbiotic importance, PRPs were a prominent component of the infection thread matrix in newly infected root cells and in nodules. PRPs were also secreted by cells in the uninfected nodule parenchyma, where they were found occluding intercellular spaces outside the middle lamella. Despite structural conservation among members of this class of cell wall proteins, PRPs were targeted to distinct layers of the extracellular matrix dependent upon cell type, and may thus play separate roles in the biology of plant cells. The putative functions and the potential for interactions between PRPs and other wall polymers are discussed.Abbreviations DTT dithiothreitol - EDTA ethylenediamine tetraacetate - GRP glycine-rich protein - PCR polymerase chain reaction - PGA polygalacturonic acid - PMSF phenylmethylsulfonyl fluoride - PRP proline-rich protein - SDS-PAGE sodium dodecylsulfate-polyacrylamide gel electrophoresis - Tris tris(hydroxylmethyl) aminomethane - Tween 20 polyoxyethylene sorbitan monolaurate Dedicated to the memory of Professor John G. Torrey