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 surface-labeled 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.
Alleman
M M
MM
Department of Immunology and Infectious Diseases, Johns Hopkins University School of Hygiene and Public Health, Baltimore, Maryland 21205.
Gottlieb
M
M
D000818:Animals; D001345:Autoradiography; D003421:Crithidia / Q000201:enzymology / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D008565:Membrane Proteins / Q000096:biosynthesis; D008970:Molecular Weight; D009708:Nucleotidases / Q000096:biosynthesis* / Q000378:metabolism; D010710:Phosphates / Q000378:metabolism*; D015800:Protozoan Proteins / Q000096:biosynthesis*; D011687:Purines / Q000378:metabolism*
D008565:Membrane Proteins; D010710:Phosphates; D015800:Protozoan Proteins; D011687:Purines; D009708:Nucleotidases; C040470:3'-nucleotidase