CRITHIDIA-LUCILIAE - STARVATION FOR PURINES AND OR PHOSPHATE LEADS TO THE ENHANCED SURFACE EXPRESSION OF A PROTEIN RESPONSIBLE FOR 3'-NUCLEOTIDASE NUCLEASE ACTIVITY

It has been shown previously that starvation of the trypanosomatid protozoan Crithidia luciliae for purines and/or inorganic phosphate results in increased levels of a surface membrane-associated 3′-nucleotidase/nuclease (3′-N'ase) activity which hydrolyzes both 3′-ribonucleotides and nucleic acids, thereby permitting the organisms to transport these essential nutrients across their cell membranes. A polypeptide with the requisite catalytic properties has been identified by an in situ gel activity assay following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). In current studies, differential synthesis of the protein responsible for the 3′-N'ase activity was not demonstrable by comparisons of SDS-PAGE patterns of nutrient-replete or purine-starved parasites metabolically labeled with either [35S]methionine, [3H]leucine, or [3H]tyrosine. However, surface labeling of nutrient-replete and purine-starved cells revealed the enhanced expression of an 125I surfacelabeled 43-kDa protein which comigrated with the 3′-N'ase activity in one- and two-dimensional electrophoretic systems. The amount of this surface-labeled peptide correlated with the level of 3′-N'ase activity as measured by test tube assay. Refeeding adenosine to purine-starved cells led to the loss of both the enzyme activity and the surface iodinatable 43-kDa band as a result of renewed cell division. Starvation of these organisms for phosphate also led to the enhanced expression of the 43-kDa radioiodinatable band. The results indicated that the 3′-N'ase protein, itself, is differentially expressed at the cell surface under conditions which lead to increased enzyme activity. © 1990.