Analysis of 5' nucleotidase and alkaline phosphatase by gene disruption in Dictyostelium.
In Dictyostelium discoideum a phosphatase with a high pH optimum is known to increase in activity during cell differentiation and become localized to a narrow band of cells at the interface of prespore and prestalk cells. However, it was not clear if this activity is due to a classical "alkaline phosphatase" with broad range substrate specificity or to a "5'nucleotidase" with high substrate preference for 5'AMP. We attempted to disrupt the genes encoding these two phosphatase activities in order to determine if the activity that is localized to the interface region resides in either of these two proteins. During aggregation of 5nt null mutants, multiple tips formed rather than the normal single tip for each aggregate. In situ phosphatase activity assays showed that the wt and the 5nt gene disruption clones had normal phosphatase activity in the area between prestalk and prespore cell types, while the alp null mutants did not have activity in this cellular region. Thus, the phosphatase activity that becomes localized to the interface of the prestalk and prespore cells is alkaline phosphatase.
Rutherford
Charles L
CL
Biology Department, Molecular and Cellular Biology Section, Virginia Polytechnic Institute and State University, 2119 Derring Hall, Blacksburg, VA 24061-0406, USA. rutherfo@vt.edu
Overall
Danielle F
DF
Ubeidat
Muatasem
M
Joyce
Bradley R
BR
D015720:5'-Nucleotidase / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D004023:Dictyostelium / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D010641:Phenotype; D016133:Polymerase Chain Reaction
D000469:Alkaline Phosphatase; D015720:5'-Nucleotidase