An unexplained sequestration of latrunculin A is required in neutrophils for inhibition of actin polymerization.
Latrunculin A (LatA) is a toxic natural product that causes disruption of the actin cytoskeleton in many eukaryotic cells at submicromolar concentrations. LatA has been found to bind G-actin with a dissociation constant of 0.2 microM, and more recently to bind profilin-G-actin and, weakly, thymosin beta4-G-actin. A number of investigators have used LatA as a G-actin sequestering agent. Thus, we studied neutrophil chemotaxis and its requisite conversion of G-actin to F-actin, supported by an extensive pool of G-actin, mainly bound to thymosin beta4. Calculations suggest that the affinity of LatA is insufficient to cause significant sequestration of this pool, and the pool's buffering action should protect neutrophils from depletion of productive G-actin species by submicromolar LatA. Nonetheless, we found that both chemoattractant stimulated migration and F-actin polymerization in neutrophils were inhibited by LatA at these concentrations. The latter effect was accompanied by sequestration of LatA and showed a cell density dependence that was consistent with G-actin sequestration. The apparent contradiction between the calculations and the experimental observations could be reconciled by assuming the presence of an accessory species, of unknown normal function, which forms a high affinity ternary complex with LatA and G-actin, thus causing the cells to concentrate LatA. Other models that could not be ruled out also invoke new actions of LatA, suggesting caution in the interpretation of its effects on cells.
Department of Physiology, University of Pennsylvania, Philadelphia 19104-6085, USA. pringm@dolphin.upenn.edu
Martin
M
Pring
Lynne
L
Cassimeris
Sally H
SH
Zigmond
D008841:Actin Cytoskeleton / Q000187:drug effects; D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism*; D000818:Animals; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D002452:Cell Count; D002633:Chemotaxis; D003470:Culture Media; D051345:Destrin; D007961:Leukocyte Migration-Inhibitory Factors / Q000378:metabolism; D008840:Microfilament Proteins / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D011817:Rabbits; D013844:Thiazoles / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D053778:Thiazolidines; D013997:Time Factors
D051339:Actin Depolymerizing Factors; D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D003470:Culture Media; D051345:Destrin; D007961:Leukocyte Migration-Inhibitory Factors; D008840:Microfilament Proteins; D013844:Thiazoles; D053778:Thiazolidines; C037067:latrunculin A