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Subunit of
the Sodium Pump Is Required for Acquisition of Fluid
Transport Capacity during Mouse Blastocyst Development
Department of Physiology, The University of Western Ontario, London, Ontario N6A 5C1
The sodium/potassium pump, Na+,K+-ATPase, is generally understood to function as a heterodimer of two subunits, a catalytic 
subunit and a
noncatalytic, glycosylated 
subunit. Recently, a putative third subunit, the 
subunit, was cloned. This small protein (6.5 kD) coimmunoprecipitates with the and
subunits and is closely associated with the ouabain
binding site on the holoenzyme, but its function is unknown. We have investigated the expression of the subunit in preimplantation mouse development, where
Na+,K+-ATPase plays a critical role as the driving
force for blastocoel formation (cavitation). Using reverse transcriptase-polymerase chain reaction, we demonstrated that the subunit mRNA accumulates continuously from the eight-cell stage onward and that it
cosediments with polyribosomes from its time of first
appearance. Confocal immunofluorescence microscopy
revealed that the subunit itself accumulates and is localized at the blastomere surfaces up to the blastocyst
stage. In contrast with the and subunits, the subunit is not concentrated in the basolateral surface of the polarized trophectoderm layer, but is strongly expressed at the apical surface as well. When embryos
were treated with antisense oligodeoxynucleotide complementary to the subunit mRNA, ouabain-sensitive K+ transport (as indicated by 86Rb+ uptake) was reduced and cavitation delayed. However, Na+,K+-ATPase enzymatic activity was unaffected as determined by
a direct phosphorylation assay ("back door" phosphorylation) applied to plasma membrane preparations.
These results indicate that the subunit, although not
an integral component of Na+,K+-ATPase, is an important determinant of active cation transport and that, as
such, its embryonic expression is essential for blastocoel formation in the mouse.
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