Tyrosine phosphates act on steroidogenesis through the activation of arachidonic acid release.
The ACTH signaling pathway includes both PKA activation as well as PKA-dependent tyrosine phosphatase activation. In addition, the action of this hormone also includes the regulation of the intracellular levels of arachidonic acid (AA) by the concerted action of two enzymes: an acylCoA-thioesterase and an acyl-CoA-synthetase (ACS4). This work describes the production and characterization of a specific ACS4 antibody, which was used to analyze the effect of ACTH on ACS4 protein level in Y1 adrenocortical cells and the putative relationship between tyrosine phosphatases and ACS4. The antiserum was obtained from rabbits immunized with the recombinant ACS4. This immunogen was produced in bacteria and eluted from an acrylamide gel after SDS-PAGE separation of a partially purified bacteria lysate. When used in Western blot analysis, the antibody obtained specifically recognized only one protein of the molecular mass corresponding to ACS4, in Y1 cells and in several rat tissues. Using the antibody described here, we analyzed the effect of ACTH stimulation on ACS4 protein level. The hormone produced an increase of this acyl-CoA synthetase in Y1 adrenocortical cells. Moreover, this effect was mimicked by cAMP and partially reduced by a tyrosine phosphatase inhibitor. We propose that ACTH regulates ACS4 protein levels through a PKA-dependent mechanism that could involve also PTP activity.
Castillo
Fernanda
F
Department of Biochemistry, School of Medicine, University of Buenos Aires, Buenos Aires, Argentina.
Cano
Florencia
F
Maloberti
Paula
P
Castilla
Rocío
R
Neuman
Isabel
I
Poderoso
Cecilia
C
Paz
Cristina
C
Podestá
Ernesto J
EJ
Cornejo Maciel
Fabiana
F
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000302:Adrenal Cortex / Q000166:cytology* / Q000378:metabolism; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D001152:Arsenicals / Q000494:pharmacology; D015153:Blotting, Western; D002460:Cell Line; D003066:Coenzyme A Ligases / Q000276:immunology / Q000378:metabolism*; D004789:Enzyme Activation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007106:Immune Sera / Q000096:biosynthesis; D008297:Male; D051379:Mice; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000276:immunology; D013256:Steroids / Q000096:biosynthesis*
D001152:Arsenicals; D004791:Enzyme Inhibitors; D007106:Immune Sera; D011994:Recombinant Proteins; D013256:Steroids; C029341:oxophenylarsine; D015124:8-Bromo Cyclic Adenosine Monophosphate; D016718:Arachidonic Acid; D000324:Adrenocorticotropic Hormone; D017027:Protein Tyrosine Phosphatases; C500958:Acsl4 protein, rat; D003066:Coenzyme A Ligases