Cell surface and organelle transport share the same enzymatic properties in Reticulomyxa.
Reticulomyxa transports particulates, like bacterial and algal prey items, bidirectionally along the outside of its pseudopodia. This cell surface transport and intracellular organelle transport can be reactivated in detergent permeabilized cell models [Orokos et al., 1997: Cell Motil. Cytoskeleton]. We have used this unique system to compare cell surface and organelle mechanochemistry in situ in the same reactivated pseudopodia. The ATPase activities of both types of transport were indistinguishable; each displayed identical nucleoside triphosphate specificity, transport ATPase kinetics, and inhibitor sensitivity. Organelle and cell surface transport reactivation required "hydrolyzable" adenosine nucleoside triphosphates; neither reactivated with GTP, CTP, UTP, ITP, AMP-PNP, AMP-PCP, or ATP-gamma-S. However, other ATP analogues, such as 2'-deoxy-ATP and 3'-deoxy-ATP and 2',3'-dideoxy-ATP, supported the reactivation of organelle and cell surface transport at similar, but markedly reduced, velocities. Both transport processes were inhibited similarly by known inhibitors of dynein ATPases such as erythro-9-(3-[2-hydroxynonyl]) adenine (EHNA) or sodium (Na)-orthovanadate. N-ethylmaleimide (NEM) and ultraviolet (UV) irradiation in the presence of Na-orthovanadate and ATP permanently disabled both transport processes. Organelle and surface transport followed identical Michaelis-Menten kinetics with a calculated Km of 118 microM ATP and a maximum translocation velocity (Vmax) of 8.33 microm/sec. These findings strongly suggest that cell surface transport shares the same cytoplasmic dynein motor [Schliwa et al., 1991: J. Cell Biol. 112:1199-1203] that drives organelle transport.
Orokos
D D
DD
Department of Biological Sciences, University of Albany, State University of New York, 12222, USA.
Travis
J L
JL
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000656:Amoeba / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D016014:Linear Models; D018715:Microscopy, Video; D015388:Organelles / Q000187:drug effects / Q000201:enzymology*; D014466:Ultraviolet Rays
D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases