Cell-cell/cell-matrix recognition phenomena are essential to numerous biological processes (normal and pathogenic). We wish to determine in detail the molecular rules governing specific macromolecular associations involved in cell-adhesion and extracellular matrix assembly. Chlamydomonas is an excellent experimental organism for such studies due to the facility of biochemical, structural, and genetic analyses. Recognition/adhesion of molecules (agglutinins) and extracellular matrix (cell wall) components have been isolated from Chlamydomonas and shown to be members of a family of hydroxyproline-rich glycoproteins (HRGP's) that may share a common evolutionary heritage. Each class of HRGP engages in specific binding activities in vitro in a manner that mimics their in vivo function. What constitutes a specific binding domain? How do these domains differ between homologous (plus and minus) agglutinin molecules? Are homologous domains responsible for self-assembly of cell wall components? Are agglutinin and matrix HRGP's members of a multigene family? cDNA libraries will be constructed from poly A+ RNA of vegetative and gametic cells of each mating type (mt+ and mt-) of Chlamydomonas, in the expression vector Lambdagtll, or in Lambdagt10. Libraries will be screened using antibody (Lambdagtll) or synthetic oligonucleotide (Lambdagt10) probes to identify inserts coding for mt+ and mt- agglutinins and GPl, a component of the crystalline layer of the cell wall that appears to be related to the agglutinins by several criteria. Candidate binding sequences for agglutinins will be identified by differential hybridization. Antibodies produced against corresponding fusion proteins, will be used for inhibition assays and epitope-mapping. Functional domains will be examined further, using synthetic peptides anti-peptide antibodies. GPl binding domains, identified in the same manner, will be mapped and compared with agglutinin sequences for evidence of homology.
| Adair, W S; Apt, K E (1990) Cell wall regeneration in Chlamydomonas: accumulation of mRNAs encoding cell wall hydroxyproline-rich glycoproteins. Proc Natl Acad Sci U S A 87:7355-9 |
| Adair, W S; Steinmetz, S A; Mattson, D M et al. (1987) Nucleated assembly of Chlamydomonas and Volvox cell walls. J Cell Biol 105:2373-82 |
