Characterization of the 70 kDa polypeptide of the Na/Ca exchanger

Saba, Rami I.; Bollen, Alex; Herchuelz, André

doi:10.1042/bj3380139

Cited by 16 publications

(20 citation statements)

References 31 publications

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“…Although the presence of a Na ϩ ͞Ca 2ϩ exchange mechanism in bovine sperm plasma membrane vesicles has been suggested (30), the present study demonstrates the presence of such an exchanger on the surface of intact sperm. Not only does the Na ϩ ͞Ca 2ϩ exchanger in herring sperm comigrate with the canine myocyte polypeptide at 120 kDa, a 70-kDa band was recognized by the antibody in some preparations, which corresponds to a proteolytic degradation product or subunit of the exchanger (31,32). Interestingly, the Na ϩ ͞Ca 2ϩ exchanger localization studies produced similar binding patterns as a study (17) that localized SMIF-binding sites over the entire sperm surface, with the majority of label over the midpiece region.…”

Section: Discussionmentioning

confidence: 99%

Motility initiation in herring sperm is regulated by reverse sodium-calcium exchange

Vines

Yoshida

Griffin

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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Sperm of the Pacific herring, Clupea pallasi, are unique in that they are immotile upon spawning in the environment. Herring sperm have evolved to remain motionless for up to several days after spawning, yet are still capable of fertilizing eggs. An egg chorion ligand termed ''sperm motility initiation factor'' (SMIF) induces motility in herring sperm and is required for fertilization. In this study, we show that SMIF induces calcium influx, sodium efflux, and a membrane depolarization in herring sperm. Sperm motility initiation by SMIF depended on decreased extracellular sodium (<350 mM) and could be induced in the absence of SMIF in very low sodium seawater. Motility initiation depended on > 1 mM extracellular calcium. Calcium influx caused by SMIF involved both the opening of voltage-gated calcium channels and reverse sodiumcalcium (Na ؉ ͞Ca 2؉ ) exchange. Membrane depolarization was slightly inhibited by a calcium channel blocker and markedly inhibited by a Na ؉ ͞Ca 2؉ exchange inhibitor. Sodium efflux caused by SMIF-initiated motility was observed when using both extracellular and intracellular sodium probes. A Na ؉ ͞Ca 2؉ exchange antigen was shown to be present on the surface of the sperm, primarily over the midpiece, by using an antibody to the canine Na ؉ ͞Ca 2؉ exchanger. This antibody recognized a 120-kDa protein that comigrated with the canine myocyte Na ؉ ͞Ca 2؉ exchanger. Sperm of Pacific herring are now shown to use reverse Na ؉ ͞Ca 2؉ exchange in motility initiation. This mechanism of regulation of motility initiation may have evolved for both maintenance of immotility after spawning as well as ligand-induced motility initiation.T eleost fish sperm are quiescent within the testes and seminal plasma before spawning, but most initiate motility after dilution into the external medium (freshwater or seawater for most species) in which spawning occurs (1). In salmonids, motility initiation occurs with dilution in freshwater, specifically from a reduction in extracellular potassium that drives a membrane hyperpolarization and an increase in intracellular calcium ([Ca 2ϩ ] i ) (2-6). Changes in the concentrations of specific ions (Ca 2ϩ , K ϩ , and possibly Na ϩ and Cl Ϫ ) also have been linked to motility initiation in Atlantic croaker sperm (7). A hyperpolarization of the sperm membrane also has been documented in carp sperm, reportedly linked to the opening of voltage-gated Ca 2ϩ channels and an increase in [Ca 2ϩ ] i (8). In other freshwater teleost sperm (goldfish, zebra fish), as well as in marine teleost sperm (puffer, flounder), motility is believed to occur as a result of nonspecific hypo-or hyperosmotic changes that drive changes in intracellular ion concentrations (5, 9, 10). Thus, in all systems studied to date, a membrane hyperpolarization leads to an increase in [Ca 2ϩ ] i and the initiation of motility.Although the stimulation of sperm motility in the vicinity of eggs has been reported in some teleost fish, only herring sperm have been shown to require an egg chorion-derived ligand for ...

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Section: Discussionmentioning

confidence: 99%

Motility initiation in herring sperm is regulated by reverse sodium-calcium exchange

Vines

Yoshida

Griffin

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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“…Apparently, in one portion of the protein, chymotrypsin cleaves the exchanger at multiple sites, and the fragments are too small to be detected in the gel system. In two studies, the N terminus of the 70-kDa proteolytic fragment has been identified to coincide with the N terminus of the full-length protein (27) or to begin within the intracellular loop in the 257-269-residue region (28).…”

Section: Split Na ϩ -Ca 2ϩ Exchangersmentioning

confidence: 99%

Split Na+-Ca2+ Exchangers

Ottolia

John

Qiu

et al. 2001

Journal of Biological Chemistry

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The Na ؉ -Ca 2؉ exchanger has nine transmembrane segments, with a large cytoplasmic loop between the fifth and sixth transmembrane segments. The protein was split within the cytoplasmic loop into two domains consisting of the first five transmembrane segments and the last four transmembrane segments, respectively. The two domains were either expressed individually or coexpressed. Each of the two domains with different lengths of the cytoplasmic loop was fused to green fluorescent protein. We show that coexpression of both domains is required for proper membrane targeting and for expression of functional exchange activity. Fusion to green fluorescent protein does not alter biophysical properties of the exchange process. In addition, truncation of a large portion of the cytoplasmic loop does not alter important properties of the exchanger such as Na ؉

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“…We used the recombinant plasmid to express V5/His-tagged NCX1.5 in HEK293 cells. Immunoblot analysis using anti-V5 antibody revealed that transfection of HEK293 cells with the NCX1.5-V5/His plasmid resulted in a single broad band in lysates with a molecular mass of 120 kDa, a component of the native full-length exchanger (9). These findings suggest that the NCX1.5-V5/His plasmid-transfected HEK293 cells highly expressed the full-length NCX1.5 tagged with the V5 epitope.…”

Section: Short Communicationmentioning

confidence: 85%

Effect of Overexpression of the Brain-Specific Na+/Ca2+ Exchanger Splice Variant NCX1.5 on NO Cytotoxicity in HEK293 Cells

et al. 2013

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Abstract. Nitric oxide (NO) induces cytotoxicity in neuronal and glial cells via activation of the Na + /Ca 2+ exchanger (NCX). This study examined the role of the predominant brain-specific NCX splice variant NCX1.5 in NO-induced cytotoxicity in the HEK293 cell expression system. Cells were transfected with the plasmid construct pcDNA3.1/V5-His containing full-length rat NCX1.5 cDNA. There was no difference in the cytotoxic effects of the NO donors sodium nitroprusside and S-nitroso-N-acetylpenicillamine between control and transfected cells. These results suggest that NO cytotoxicity is not dependent on NCX1.5.

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Characterization of the 70 kDa polypeptide of the Na/Ca exchanger

Cited by 16 publications

References 31 publications

Motility initiation in herring sperm is regulated by reverse sodium-calcium exchange

Motility initiation in herring sperm is regulated by reverse sodium-calcium exchange

Split Na+-Ca2+ Exchangers

Effect of Overexpression of the Brain-Specific Na+/Ca2+ Exchanger Splice Variant NCX1.5 on NO Cytotoxicity in HEK293 Cells

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