Post-transcriptional Control of Na,K-ATPase Activity and Cell Growth by a Splice Variant of FXYD2 Protein with Modified mRNA

被引:7
作者
Sweadner, Kathleen J.
Pascoa, Jennifer L.
Salazar, Cynthia A.
Arystarkhova, Elena [1 ]
机构
[1] Massachusetts Gen Hosp, Dept Neurosurg, Lab Membrane Biol, Boston, MA 02114 USA
基金
美国国家卫生研究院;
关键词
ION-TRANSPORT REGULATORS; SODIUM-POTASSIUM PUMP; NA+-K+-ATPASE; GAMMA-SUBUNIT; ENDOPLASMIC-RETICULUM; RENAL NA; K-ATPASE; TRANSMEMBRANE PROTEINS; CRYSTAL-STRUCTURE; MAMMALIAN-CELLS; KINASE-C;
D O I
10.1074/jbc.M111.241901
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
In kidney, FXYD proteins regulate Na,K-ATPase in a nephron segment-specific way. FXYD2 is the most abundant renal FXYD but is not expressed in most renal cell lines unless induced by hypertonicity. Expression by transfection of FXYD2a or FXYD2b splice variants in NRK-52E cells reduces the apparent Na+ affinity of the Na, K-ATPase and slows the cell proliferation rate. Based on RT-PCR, mRNAs for both splice variants were expressed in wild type NRK-52E cells as low abundance species. DNA sequencing of the PCR products revealed a base alteration from C to T in FXYD2b but not FXYD2a from both untreated and hypertonicity-treated NRK-52E cells. The 172C -> T sequence change exposed a cryptic KKXX endoplasmic reticulum retrieval signal via a premature stop codon. The truncation affected trafficking of FXYD2b and its association with Na,K-ATPase and blocked its effect on enzyme kinetics and cell growth. The data may be explained by altered splicing or selective RNA editing of FXYD2b, a supplementary process that would ensure that it was inactive even if transcribed and translated, in these cells that normally express only FXYD2a. 172C -> T mutation was also identified after mutagenesis of FXYD2b by error-prone PCR coupled with a selection for cell proliferation. Furthermore, the error-prone PCR alone introduced the mutation with high frequency, implying a structural peculiarity. The data confirm truncation of FXYD2b as a potential mechanism to regulate the amount of FXYD2 at the cell surface to control activity of Na,K-ATPase and cell growth.
引用
收藏
页码:18290 / 18300
页数:11
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