IMMUNOCYTOCHEMICAL LOCALIZATION OF PHENYLALANINE AMMONIA-LYASE IN TISSUES OF POPULUS-KITAKAMIENSIS

The polypeptide encoded by the partial fragment of cDNA of phenylalani ne ammonia-lyase (PAL; EC 4.3.1.5), PALcDNA1 (Osakabe et al., 1995, Pl ant Sci. 105: 217-226), isolated from Populus kitakamiensis (P. siebol dii x P. grandidentata), was expressed in Escherichia coli cells. The polypeptide was purified and an antiserum raised against it. The antis erum recognized a protein of 77 kDa on nitrocellulose blots after sodi um dodecyl sulfate-polyacrylamide gel electrophoresis of total protein and the partially purified PAL protein from P. kitakamiensis. Moreove r,the antiserum recognized a protein on the blot after non-denaturing polyacrylamide gel electrophoresis of P. kitakamiensis proteins and th is protein had PAL activity. Furthermore, the antibody inhibited PAL a ctivity of extracts from stem tissues. These results showed that the a ntiserum against the partial PAL peptide recognized only the PAL subun its in extracts of P. kitakamiensis. Immunolocalization studies of P. kitakamiensis tissues revealed that the PAL protein was specifically l ocalized in the xylem and the phloem fibers and no immunogold signal w as found in the epidermis, the cortex, the pith, or the cambium of eit her stems or leaves.