Constitutively active protein kinase A disrupts motility and chemotaxis in Dictyostelium discoideum.
The deletion of the gene for the regulatory subunit of protein kinase A (PKA) results in constitutively active PKA in the pkaR mutant. To investigate the role of PKA in the basic motile behavior and chemotaxis of Dictyostelium discoideum, pkaR mutant cells were subjected to computer-assisted two- and three-dimensional motion analysis. pkaR mutant cells crawled at only half the speed of wild-type cells in buffer, chemotaxed in spatial gradients of cyclic AMP (cAMP) but with reduced efficiency, were incapable of suppressing lateral pseudopods in the front of temporal waves of cAMP, a requirement for natural chemotaxis, did not exhibit the normal velocity surge in response to the front of a wave, and were incapable of chemotaxing toward an aggregation center in natural waves generated by wild-type cells that made up the majority of cells in mixed cultures. Many of the behavioral defects appeared to be the result of the constitutively ovoid shape of the pkaR mutant cells, which forced the dominant pseudopod off the substratum and to the top of the cell body. The behavioral abnormalities that pkaR mutant cells shared with regA mutant cells are discussed by considering the pathway ERK2 perpendicular RegA perpendicular [cAMP] --> PKA, which emanates from the front of a wave. The results demonstrate that cells must suppress PKA activity in order to elongate along a substratum, suppress lateral-pseudopod formation, and crawl and chemotax efficiently. The results also implicate PKA activation in dismantling cell polarity at the peak and in the back of a natural cAMP wave.
Zhang
Hui
H
W M Keck Dynamic Image Analysis Facility, Department of Biological Sciences, University of Iowa, Iowa City, Iowa 52242, USA.
Heid
Paul J
PJ
Wessels
Deborah
D
Daniels
Karla J
KJ
Pham
Tien
T
Loomis
William F
WF
Soll
David R
DR
D015105:3',5'-Cyclic-AMP Phosphodiesterases; D000818:Animals; D016764:Cell Polarity / Q000187:drug effects / Q000235:genetics; D048429:Cell Size / Q000187:drug effects / Q000235:genetics; D002633:Chemotaxis / Q000187:drug effects / Q000235:genetics*; D000242:Cyclic AMP / Q000378:metabolism / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D004023:Dictyostelium / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D007091:Image Processing, Computer-Assisted; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D011554:Pseudopodia / Q000201:enzymology* / Q000235:genetics / Q000648:ultrastructure
D015800:Protozoan Proteins; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D015105:3',5'-Cyclic-AMP Phosphodiesterases; C112742:regA protein, Dictyostelium