Na,K-ATPase from Mice Lacking the γ Subunit (FXYD2) Exhibits Altered Na+ Affinity and Decreased Thermal Stability

Abstract

The ␥ subunit of the Na,K-ATPase, a 7-kDa single-span membrane protein, is a member of the FXYD gene family. Several FXYD proteins have been shown to bind to Na,K-ATPase and modulate its properties, and each FXYD protein appears to alter enzyme kinetics differently. Different results have sometimes been obtained with different experimental systems, however. To test for effects of ␥ in a native tissue environment, mice lacking a functional ␥ subunit gene (Fxyd2) were generated. These mice were viable and without observable pathology. Prior work in the mouse embryo showed that ␥ is expressed at the blastocyst stage. However, there was no delay in blastocele formation, and the expected Mendelian ratios of offspring were obtained even with Fxyd2 ؊/؊ dams. In adult Fxyd2 ؊/؊ mouse kidney, splice variants of ␥ that have different nephron segment-specific expression patterns were absent. Purified ␥-deficient renal Na,K-ATPase displayed higher apparent affinity for Na ؉ without significant change in apparent affinity for K ؉ . Affinity for ATP, which was expected to be decreased, was instead slightly increased. The results suggest that regulation of Na ؉ sensitivity is a major functional role for this protein, whereas regulation of ATP affinity may be context-specific. Most importantly, this implies that ␥ and other FXYD proteins have their effects by local and not global conformation change. Na,K-ATPase lacking the ␥ subunit had increased thermal lability. Combined with other evidence that ␥ participates in an early step of thermal denaturation, this indicates that FXYD proteins may play an important structural role in the enzyme complex.

Inherited forms of renal hypomagnesemia: an update

“…These findings strongly suggest that the hypomagnesemia in the patients with FXYD2 mutations is caused by a dominant negative effect rather than by haploinsufficiency. The recent finding that Fxyd2 knockout mice have normal serum Mg 2+ levels and do not show any abnormalities in Mg 2+ excretion is consistent with this assumption [46].…”

Structural Interactions between FXYD Proteins and Na+,K+-ATPase

“…By contrast, however, transfection of ␥ into Xenopus oocytes or HeLa cells is not associated with significant effects on V max values (13,14). In renal microsomes isolated from ␥ knock-out mice, the major effect appears to be that on the cytoplasmic Na ϩ affinity (16). CHIF raises the apparent affinity for cell Na ϩ by 2-3-fold, the reverse effect to that of ␥ (13,17).…”

Grainyhead-related transcription factor is required for duct maturation in the salivary gland and the kidney of the mouse

“…In addition, some other genes also failed to be expressed in the SMG duct of Cp2l1 tra/tra mice. The expression of Gk-6 (Penschow et al, 1991), Fxyd2a and Fxyd2b (genes encoding Na + ,K + -ATPase ␥-subunit variants a and b) (Jones et al, 2001;Jones et al, 2005), and Scnn1b (gene encoding the ␤-subunit of the epithelial sodium channel) in the SMG was detected during normal development but severely reduced in Cp2l1 tra/tra embryos throughout development (Fig. 3B-D,F and data not shown).…”