title: Oilseed isocitrate lyases lacking their essential type 1 peroxisomal targeting signal are piggybacked to glyoxysomes.
abstract: Isocitrate lyase (IL) is an essential enzyme in the glyoxylate cycle, which is a pathway involved in the mobilization of stored lipids during postgerminative growth of oil-rich seedlings. We determined experimentally the necessary and sufficient peroxisome targeting signals (PTSs) for cottonseed, oilseed rape, and castor bean ILs in a well-characterized in vivo import system, namely, suspension-cultured tobacco (Bright Yellow) BY-2 cells. Results were obtained by comparing immunofluorescence localizations of wild-type and C-terminal-truncated proteins transiently expressed from cDNAs introduced by microprojectile bombardment. The tripeptides ARM-COOH (on cottonseed and castor bean ILs) and SRM-COOH (on oilseed rape IL) were necessary for targeting and actual import of these ILs into glyoxysomes, and ARM-COOH was sufficient for redirecting chloramphenicol acetyltransferase (CAT) from the cytosol into the glyoxysomes. Surprisingly, IL and CAT subunits without these tripeptides were still acquired by glyoxysomes, but only when wild-type IL or CAT-SKL subunits, respectively, were simultaneously expressed in the cells. These results reveal that targeting signal-depleted subunits are being piggybacked as multimers to glyoxysomes by association with subunits possessing a PTS1. Targeted multimers are then translocated through membrane pores or channels to the matrix as oligomers or as subunits before reoligomerization in the matrix.
authors.affiliation: Department of Botany, Arizona State University, Tempe 85287-1601, USA.
authors.forename: M S
authors.initials: MS
authors.lastname: Lee
authors.forename: R T
authors.initials: RT
authors.lastname: Mullen
authors.forename: R N
authors.initials: RN
authors.lastname: Trelease
mesh_terms: D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003369:Cottonseed Oil; D018076:DNA, Complementary / Q000737:chemistry; D007887:Fabaceae; D007522:Isocitrate Lyase / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008830:Microbodies / Q000201:enzymology / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D010946:Plants, Medicinal; D010947:Plants, Toxic; D011956:Receptors, Cell Surface / Q000378:metabolism*; D014026:Nicotiana
chemical_list: D003369:Cottonseed Oil; D018076:DNA, Complementary; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; C085380:peroxisomal targeting sequence receptor; D015500:Chloramphenicol O-Acetyltransferase; D007522:Isocitrate Lyase