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EDITORIAL FOCUS

The stoichiometry of the Na-K pump: one plus one doesn't equal one

Department of Cell Physiology and Molecular Biophysics, Texas Tech University Health Sciences Center, Lubbock, Texas

PICK UP NEARLY ANY TEXTBOOK of cell physiology, and you will read that the constituent subunits of the Na-K pump exist in equimolar amounts. As a central player in Na⁺ and K⁺ transport, this protein complex has been the subject of intense biochemical scrutiny for decades, and the argument for a 1:1 ratio between the catalytic -subunit and its glycosylated β-subunit dates from the earliest efforts at purification of the enzyme complex. The idea that one and one β make a functional heterodimer has thus approached the status of dogma. This assertion has now been challenged in a provocative report from Clifford and Kaplan (1), who argue that, under conditions of subunit overexpression, the pump complex may vary in its subunit stoichiometry. It appears that, with respect to the Na-K pump, one plus one really doesn't equal one.

These Chicago-based investigators used a heterologous expression strategy in Madin-Darby canine kidney cells, a longstanding culture model of renal epithelia. An inducible promoter allowed them to force overexpression of exogenous β-subunit in the recipient cells, sometimes by as much as 10-fold. Their conclusion of varying stoichiometry is based on two key results in this system. First, despite the presence of "excess" β-subunit, they observed no change in abundance of the -subunit, nor the specific activity of the enzyme. Second, they found that >95% of the expressed β-subunit was associated with . With no change in apparent pump abundance and little "free" β, Clifford and Kaplan (1) drew the obvious inference that multiple β-subunits must be associating with in the pump complex, an interpretation supported by subsequent measurements of subunit ratio. Indeed, their data suggest a stoichiometry as high as 5:1, prompting them to dismiss direct binding of multiple β-subunits to a single as an explanation. Instead, they favor a series of β-β interactions that is, in turn, anchored to a given , perhaps producing a chain of β-subunits. Biochemists have long believed that the stoichiometry of the complex was inherent in the properties and structures of the constituent subunits. These new findings suggest that this view may be too conservative, and changes in subunit expression or abundance may also play a role. If so, this places increased pressure on the regulation of subunit abundance if the composition of the pump complex is to be controlled.

The results reported by the Chicago team raise a host of new questions. Most importantly, it remains unclear if the variable stoichiometry is biologically relevant. After all, it could simply be a consequence of heterologous expression and the strong promoter that ensured an excessive number of β-subunits. Even if observed only in culture, however, the details of the subunit interactions must be identified and reconciled with the high-resolution model proposed for the pump complex from pig kidney (5). It is all the more frustrating that, other than the transmembrane domain, the remaining parts of the β-subunit are poorly resolved in the proposed structure, leaving us with few clues as to how multiple β-subunits might interact. Nevertheless, inhibition of glycosylation had no effect on the changes in stoichiometry, indicating that the N-linked glycans of β are not relevant to association. Of course, the results from Clifford and Kaplan (1) do not rule out the possibility of an additional polypeptide that could mediate this interaction. Such a situation might prove to be an embarrassment of riches, with the pump complex becoming more and more crowded with players.

If variable stoichiometry is more generally true in biology, however, one can imagine how the ratio between subunits might be regulated. Indeed, changes in gene expression of the β-subunit or its degradation induced by hormones or second messengers could allow fine tuning of the complex. But to what end? We might ask whether a changing stoichiometry has functional consequences. Indeed, it is clear that the β-subunit plays a role in catalysis (2), and it would not be farfetched if its influence varied with subunit ratio. Clifford and Kaplan (1) saw no obvious enzymatic differences, but they documented a more rapid development of transepithelial resistance in the overexpressing cells that suggests a role for the β-subunits in adhesion. A more detailed study of kinetics and function is clearly needed. Excess β may lead to βⁿ complexes, but, if there is excess , do we see corresponding ⁿβ? The few examples in the literature suggest that we do not (3, 4, 6), but a more explicit examination is probably warranted. Does this variable stoichiometry extend to other subunit isoforms? This too might yield an embarrassingly large variety of Na-K pump complexes. It seems likely that, regardless of how biologically relevant the results prove to be, Clifford and Kaplan's report (1) will send many of us back to the bench to explore further the interactions of the pump's subunits.

	FOOTNOTES

 

Address for reprint requests and other correspondence: T. A. Pressley, Dept. of Cell Physiology and Molecular Biophysics, Texas Tech Univ. Health Sciences Center, Lubbock, TX 79430 (e-mail: Thomas.Pressley{at}ttuhsc.edu)

	REFERENCES

TOP REFERENCES

 

1. Clifford RJ, Kaplan JH. β-Subunit overexpression alters the stoichiometry of assembled Na-K-ATPase subunits in MDCK cells. Am J Physiol Renal Physiol (First published August 13, 2008). doi:10.1152/ajprenal.90406.2008.[Abstract/Free Full Text]
2. Dempski RE, Friedrich T, Bamberg E. The β subunit of the Na⁺/K⁺-ATPase follows the conformational state of the holoenzyme. J Gen Physiol 125: 505–520, 2005.[Abstract/Free Full Text]
3. Factor P, Senne C, Dumasius V, Ridge K, Jaffe HA, Uhal B, Gao Z, Sznajder JI. Overexpression of the Na⁺,K⁺-ATPase 1 subunit increases Na⁺,K⁺-ATPase function in A569 cells. Am J Respir Cell Mol Biol 18: 741–749, 1998.[Abstract/Free Full Text]
4. Gomes P, Soares-da-Silva P. Upregulation of apical NHE3 in renal OK cells overexpressing the rodent ₁-subunit of the Na⁺ pump. Am J Physiol Regul Integr Comp Physiol 290: R1142–R1150, 2006.[Abstract/Free Full Text]
5. Morth JP, Pedersen BP, Toustrup-Jensen MS, Sorensen TLM, Petersen J, Andersen JP, Vilsen B, Nissen P. Crystal structure of the sodium-potassium pump. Nature 450: 1043–1049, 2007.[CrossRef][Web of Science][Medline]
6. Shanbaky NM, Pressley TA. Transfection of Na,K-ATPase -subunit: regulation of enzyme abundance. Biochem Cell Biol 73: 261–268, 1995.[Web of Science][Medline]

This Article Full Text (PDF) All Versions of this Article: 295/5/F1313    most recent 90512.2008v1 Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Pressley, T. A. Search for Related Content PubMed PubMed Citation Articles by Pressley, T. A.