Synthesis of Na+/K+ ATPase by the preimplantation rabbit blastocyst.
The rates of incorporation of [35S]methionine into Na+/K+ ATPase, actin (beta- and gamma-isoforms), and total protein of the preimplantation rabbit blastocyst were determined between Days 4 and 7 of development. Blastocyst proteins were metabolically radiolabelled with [35S]methionine and subsequently analysed by co-isolation with purified Na+/K+ ATPase using two-dimensional polyacrylamide gel electrophoresis, immunoprecipitation, immunoblotting, fluorography, and liquid scintillation spectroscopy. The rate of [35S]methionine incorporation into acid-soluble total protein increased 24-fold between Days 4 and 6 post coitum (p.c.), then diminished approximately 79% on Day 7. In-vitro incorporation of [35S]methionine was linear at each stage of blastocyst development. [35S]methionine incorporation rates were unaffected by low free intracellular methionine concentration (less than 0.06 mM) and stage-related differences in blastocoele volume. Analysis of beta- and gamma-actin synthesis revealed patterns of [35S]methionine incorporation rates which were similar to those of total protein. In contrast, synthesis of blastocyst Na+/K+ ATPase was characterized by a 90-fold increase (P less than 0.001) in the rate of [35S]methionine incorporation between Days 4 and 6 p.c. The results demonstrate that Na+/K+ ATPase is actively synthesized at a high and increasing rate during preimplantation development in the rabbit at a period which is characterized by rapid fluid accumulation by the blastocyst.
Overström
E W
EW
Department of Physiology and Biophysics, Harvard Medical School, Boston, Massachusetts 02115.
Benos
D J
DJ
Biggers
J D
JD
D000818:Animals; D001755:Blastocyst / Q000201:enzymology* / Q000378:metabolism; D005260:Female; D008715:Methionine / Q000378:metabolism; D011817:Rabbits; D000254:Sodium-Potassium-Exchanging ATPase / Q000096:biosynthesis*
D008715:Methionine; D000254:Sodium-Potassium-Exchanging ATPase