A polymorphic, prespore-specific cell surface glycoprotein is present in the extracellular matrix of Dictyostelium discoideum.

Abstract

Polymorphisms of a major developmentally regulated prespore-specific protein (PsA) in Dictyostelium discoideum slugs are described. These polymorphisms allowed discrimination between PsA (found on the cell surface and in the extracellular matrix) and a similar extracellular but nonpolymorphic protein, ShA. The two proteins were also distinguished by their differing reactivities with a range of monoclonal antibodies and by their sensitivity to release from the sheath with cellulase. The results are discussed in terms of the molecular and genetic relationships between the cell surface and the extracellular matrix during development.Interactions between the cell surface and the extracellular matrix have been proposed to play an important role in developmental processes (e.g., sea urchin blastulation [6,10]). In some cases (e.g., fibronectin), there are distinct cellular and extracellular forms of the molecules proposed to be involved in these interactions, raising questions concerning the structural and functional relationships between the cell surface and the extracellular matrix. Consequently, elucidation of the biochemical similarities and differences between cellular and extracellular molecules is a necessary prerequisite to resolve how such interactions occur.The cellular slime mold Dictyostelium discoideum has been used extensively as a model system for the study of a wide range of developmental processes (see reference 9). During development, individual vegetative amoebae aggregate to form a multicellular mass which is then covered throughout the developmental process by a thin celluloseprotein sheath or extracellular matrix (16, 17). Although numerous roles for the sheath have been suggested, evaluation of these proposals has been hampered by the lack of information concerning the composition of the sheath. It contains cellulose fibrils (1) in a highly proteinaceous matrix (5, 18), and recent work with monoclonal antibodies (mAbs) has demonstrated shared antigenic determinants on a large number of prespore cell surface and extracellular matrix proteins (3; Grant et al., manuscript in preparation). Furthermore, there are a small number of specific cell surface proteins which are also apparently in the sheath, i.e., have cellular and extracellular forms.The most prominent of these was a 32-kilodalton (kDa) antigen whose cellular and extracellular forms were immunochemically distinguishable. It was not determined whether this was due to (i) alternate processing pathways of a single precursor, (ii) similar processing of unrelated precursors, or (iii) the existence of related but genetically distinct proteins.In this work we demonstrate, using genetic (13, 22), immunochemical, and biochemical criteria, that there are two 32-kDa proteins: one (termed PsA) which is common to * Corresponding author.

Structural characterization of Dictyostelium discoideum prespore-specific gene D19 and of its product, cell surface glycoprotein PsA.

“…PsA and D. discoideum contact sites A (CsA) are glycoproteins which show major decreases in size in modB mutants (11, 44; Alexander et al, in press). The primary defect caused by the modB mutation is thought to be in the mechanism responsible for the retardation of the protein until type 2 glycosylation occurs (15 12 kDa of the molecular mass of CsA (11) and for 7 kDa of the PsA apparent molecular mass as determined by SDS-PAGE (30 kDa in NC4 and 23 kDa in the modB mutant of NC4, HU2428; data not shown). Therefore, in PsA the modB-sensitive carbohydrate accounts for 7 kDa of the 12-kDa difference between the molecular mass predicted from the translated D19 gene and the apparent molecular mass determined by SDS-PAGE.…”

“…PsA was originally defined as a 30-kilodalton (kDa) prespore-specific cell surface protein carrying a unique epitope which is recognized by MUD1 (19). By using genetic, immunological, and biochemical criteria, it was demonstrated that PsA is encoded by a single locus, pspA, and that the gene product is a protein with a molecular weight polymorphism in several wild isolates (12). Another epitope on PsA is recognized by the MAb MUD50 (13), which, in addition to PsA, recognizes a diverse group of prespore proteins (S. Alexander, E. Smith, L. Davis, A. Gooley, S. B. Por, L. Browne, and K. L. Williams, Differentiation, in press).…”

Immunodominant carbohydrate determinants in the multicellular stages of Dictyostelium discoideum

“…MUD50 and MUD62 were raised as antibodies to sheath (extracellular matrix) glycoproteins (Grant & Williams, 1983;Grant et al, 1985). Both MUD50 and MUD62 recognize patterns of glycoproteins on Western blots of sheath proteins that differ from slug proteins.…”

“…It is necessary here to clarify the situation with respect to previously published information on MUD52/62. On Western blots of slug antigens, MUD52 was originally found to recognize several glycoproteins including a 30 kDa species (Grant & Williams, 1983;Grant et al, 1985). It would now appear that this was incorrect, and that the original antibody used may have been contaminated by another antibody, MUD51.…”

“…4). Although PsA and PsB exhibit size polymorphisms in different wild strains of D. discoideum (Grant et al, 1985;Smith et al, 1989), the wild-type strain in Fig. 4 is X22, the parent strain of HU2470 and HU2471.…”