Membrane potential and amino acid transport in a mutant Chinese hamster ovary cell line.
The bioenergetics of amino acid transport system A was studied in two Chinese hamster ovary (CHO) cell lines, the parent line CHO-PEOT/1 and CHY-1, a mutant of the former exhibiting a low activity of the same transport system. The steady-state transmembrane distribution ratio of the cationic amino acid L-arginine (RARG) was employed as an indicator of membrane potential (delta psi). Evidence for the reliability of RARG to measure delta psi can be summarized as follows: (1) L-arginine transmembrane distribution increased under conditions of cell hyperpolarization and decreased under conditions of cell depolarization; (2) L-arginine distribution conformed closely to that expected for a probe of delta psi in conditions in which delta psi depends largely on the transmembrane potassium gradient; and (3) the value of delta psi obtained through a valinomycin null point experiment (-72.7 mV) was very similar to the value calculated from L-arginine distribution using the Nernst equation (-73.4 mV). The transmembrane gradient of sodium electrochemical potential (delta mu Na), the driving force for the operation of system A, was slightly higher in the mutant cell line CHY-1. In the same line, the intracellular level of the specific system A substrate MeAIB at steady state was also higher. Studies of the rheogenicity of system A in the two lines indicated that the depolarization associated with the entry of substrates of system A was proportional to the amount of amino acid taken up by the cells. Kinetic analysis showed that the low activity of system A in the mutant cell line was referrable to a decrease in transport Vmax. It is concluded that neither a decrease in energy available for the operation of system A nor a decreased efficiency of coupling of the system to delta psi is responsible for the defect observed in the mutant line.
Rotoli
B M
BM
Istituto di Patologia Generale, Università di Parma, Italy.
Bussolati
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O
Dall'Asta
V
V
Gazzola
G C
GC
D000409:Alanine / Q000378:metabolism; D000596:Amino Acids / Q000493:pharmacokinetics* / Q000494:pharmacology; D000818:Animals; D001120:Arginine / Q000378:metabolism; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D004553:Electric Conductivity / Q000187:drug effects / Q000502:physiology; D005260:Female; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology*; D010053:Ovary / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D011392:Proline / Q000378:metabolism / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology
D000596:Amino Acids; D001120:Arginine; D011392:Proline; D012964:Sodium; D000409:Alanine