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J S Trimmer

Publications and source records attributed to J S Trimmer.

65 records · Page 4Linked to original sources
PubMed1989

Primary structure and functional expression of a mammalian skeletal muscle sodium channel.

We describe the isolation and characterization of a cDNA encoding the alpha subunit of a new voltage-sensitive sodium channel, microI, from rat skeletal muscle. The 1840 amino acid microI peptide is homologous to alpha subunits from rat brain, but, like the protein from eel electroplax, lacks an extended (approximately 200) amino acid segment between homologous domains I and II. Northern blot analysis indicates that the 8.5 kb microI transcript is preferentially expressed in skeletal muscle. Sodium channels expressed in Xenopus oocytes from synthetic RNA encoding microI are blocked by tetrodotoxin and mu-conotoxin at concentrations near 5 nM. The expressed sodium channels have gating kinetics similar to the native channels in rat muscle fibers, except that inactivation occurs more slowly.

Amino Acid Sequence↗
PubMed1988

Monoclonal antibodies induce the translocation, patching, and shedding of surface antigens of sea urchin spermatozoa.

Monoclonal antibodies reacting with external domains of plasma membrane proteins of sea urchin spermatozoa cause the reversible aggregation of sperm. The rate of disaggregation is temperature dependent indicating the involvement of membrane fluidity. Immunofluorescence shows that disaggregation is temporally correlated with the movement of surface-bound antibody into large aggregates which are then shed from the cell surface. Electrophoretic analysis of the shed antigens shows that they resemble the total complement of proteins of isolated sperm membranes. The translocation and shedding of surface-bound ligands may be a common property of eukaryotic flagellar membranes.

Animals↗
PubMed1988

Monoclonal antibodies to a membrane glycoprotein induce the phosphorylation of histone H1 in sea urchin spermatozoa.

Two groups of mAbs reacting with external domains of a major sea urchin sperm membrane glycoprotein of 210 kD were isolated. Previous studies have shown that group I mAbs inhibit the acrosome reaction induced by egg jelly and also cause large increases in intracellular Ca2+ [( Ca2+]i). Group II mAbs, at comparable levels of cell surface binding, neither inhibit the egg jelly-induced acrosome reaction nor cause increases in [Ca2+]i. In this paper, we investigate the ability of these mAbs to induce the cAMP-dependent phosphorylation of sperm histone H1. Group I mAbs induce H1 phosphorylation to the same level and on the same peptide, as occurs upon treatment of sperm with egg jelly. These mAbs also activate adenylate cyclase to the same extent as egg jelly. Group II mAbs do not induce H1 phosphorylation and are only poor activators of adenylate cyclase. Group I mAbs compete with each other, but not with group II mAbs, for binding to the cell surface. These data indicate that the activation of adenylate cyclase is an initial event in the pathway leading from the binding of mAbs to a specific domain of the 210-kD protein at the cell surface, to the discrete phosphorylation of histone H1 in highly condensed sperm chromatin. The domain on the 210-kD protein recognized by group I mAbs plays a critical role in signal transduction during the early events of fertilization.

Acrosome↗
PubMed1987

Changing localizations of site-specific surface antigens during sea urchin spermiogenesis.

Whole mount preparations of dissociated testicular cells from the sea urchin, Strongylocentrotus purpuratus, were exposed to monoclonal antibodies (mAbs) directed against sperm surface proteins. Indirect immunofluorescence microscopy and Western immunoblot analysis show that mAb J18/29 binds to the entire surface of the mature spermatozoon and membrane proteins ranging in relative molecular masses from 25 to 340 kDa. MAb J18/2 binds to the acrosomal and tail regions of the mature spermatozoon and mainly to a 210-kDa membrane protein. MAb J17/30 binds to the midpiece and tail regions and monospecifically to a 60-kDa membrane protein. MAb J16/33 binds specifically to the sperm midpiece but does not bind to Western immunoblots of sperm membrane proteins. With the exception of J16/33, which shows a punctate binding pattern, all of these mAbs show uniform binding over the entire surface of the early spermatid. This uniform and complete surface binding is observed through all stages of spermiogenesis for mAb J18/29. By the midspermatid stage, when tail formation first begins, but before the nucleus condenses and the cytoplasm decreases in volume, localized binding patterns of mAbs J17/30 and J16/33 become evident. Localized binding of mAb J18/2 is not observed until the late spermatid stage. These results show that the sea urchin sperm surface is composed of at least four different domains and provide the first insight into differentiation of the cell surface during sea urchin spermatogenesis.

Animals↗
PubMed1987

Dispersal of sperm surface antigens in the plasma membranes of polyspermically fertilized sea urchin eggs.

Polyspermically fertilized Strongylocentrotus purpuratus eggs were fixed at varying times after insemination and exposed to a monoclonal antibody (mAb J18/29) directed against a group of sperm surface antigens. Indirect immunofluorescence microscopy reveals that the sperm surface components recognized by mAb J18/29 are quickly incorporated into the egg plasma membrane and begin to disperse as early as 1.5 min after insemination. At subsequent times after insemination, they undergo further dispersal so that by 45 min they are distributed evenly over the entire surface of the egg. These results provide evidence for the free lateral mobility of sperm membrane components in the fertilized egg.

Animals↗
PubMed1987

Characterization of a monoclonal antibody that induces the acrosome reaction of sea urchin sperm.

A monoclonal antibody, J18/29, induces the acrosome reaction (AR) in spermatozoa of the sea urchin Strongylocentrotus purpuratus. J18/29 induces increases in both intracellular Ca2+ and intracellular pH similar to those occurring upon induction of the AR by the natural inducer, the fucose sulfate-rich glycoconjugate of egg jelly. Lowering the Ca2+ concentration or the pH of the seawater inhibits the J18/29-induced AR, as does treatment with Co2+, an inhibitor of Ca2+ channels. The J18/29-induced AR is also inhibited by verapamil, tetraethylammonium chloride, and elevated K+. All these treatments cause similar inhibition of the egg jelly-induced AR. J18/29 reacts with a group of membrane proteins ranging in molecular mass from 340 to 25 kD, as shown by immunoprecipitation of lysates of 125I-labeled sperm and Western blots. The most prominent reacting proteins are of molecular masses of 320, 240, 170, and 58 kD. The basis of the multiple reactivity appears to reside in the polypeptide chains of these proteins, as J18/29 binding is sensitive to protease digestion but resistant to periodate oxidation. There are approximately 570,000 sites per cell for J18/29 binding. J18/29 is the only reagent of known binding specificity that induces the AR; it identifies a subset of sperm membrane proteins whose individual characterization may lead to the isolation of the receptors involved in the triggering of the AR at fertilization.

Acrosome↗
PubMed1986

Monoclonal antibodies increase intracellular Ca2+ in sea urchin spermatozoa.

Changes in intracellular free Ca2+ ([Ca2+]i) of sea urchin (Strongylocentrotus purpuratus) spermatozoa were measured using the fluorescent Ca2+ indicators fura-2 and indo-1. The intracellular pH (pHi) of sperm was also determined. The fucose sulfate-rich glycoconjugate component of egg jelly induced increases in [Ca2+]i and pHi in sperm and induced the acrosome reaction. Monoclonal antibodies (mAbs) to external domains of a 210-kDa glycoprotein of the sperm plasma membrane induced a 23-fold increase in [Ca2+]i (vs. 9-fold for fucose sulfate-rich glycoconjugate), but the mAbs did not cause the pHi to increase and did not induce the acrosome reaction. When the mAb treatment which induced an increase in [Ca2+]i was combined with an NH4Cl treatment, which increased the pHi, the acrosome reaction was induced. mAb-induced increases in [Ca2+]i were dependent on millimolar concentrations of extracellular Ca2+ and were reversed by placing sperm in Ca2+-free seawater or by chelating Ca2+ with EGTA. The mAb-induced [Ca2+]i increase was sensitive to the pH of the seawater, although mAb binding was not. The data show that increased [Ca2+]i and pHi are necessary for induction of the acrosome reaction and suggest that the 210-kDa protein may play a role in regulating Ca2+ entry into the spermatozoan. These mAbs make it possible to separate the increase in [Ca2+]i from the increase in pHi and may be useful in the elucidation of the regulatory role of Ca2+ in sperm physiology.

Acrosome↗
PubMed1985

Monoclonal antibody to a membrane glycoprotein inhibits the acrosome reaction and associated Ca2+ and H+ fluxes of sea urchin sperm.

Monoclonal antibodies were prepared using sea urchin (Strongylocentrotus purpuratus) sperm or isolated sperm plasma membrane vesicles as immunogen. Two monoclonal antibodies, J4/4 and J10/14, react with different epitopes on external domains of the same Mr = 210,000 integral-membrane glycoprotein. At Fab fragment concentrations of equivalent cell surface binding, J10/14 inhibits the egg jelly-induced exocytotic acrosome reaction and associated Ca2+ influx and H+ efflux, whereas J4/4 does not. Immunofluorescent localization shows that both monoclonal antibodies react with a narrow collar of plasma membrane over the acrosomal complex and also with the entire flagellum.

Acrosome↗
PubMedDate not supplied

K+ channel distribution and clustering in developing and hypomyelinated axons of the optic nerve.

The localization of Shaker-type K(+) channels in specialized domains of myelinated central nervous system axons was studied during development of the optic nerve. In adult rats Kv1.1, Kv1.2, Kv1.6, and the cytoplasmic beta-subunit Kvbeta2 were colocalized in juxtaparanodal zones. During development, clustering of K(+) channels lagged behind that for nodal Na(+) channels by about 5 days. In contrast to the PNS, K(+) channels were initially expressed fully segregated from nodes and paranodes, the latter identified by immunofluorescence of Caspr, a component of axoglial junctions. Clusters of K(+) channels were first detected at postnatal day 14 (P14) at a limited number of sites. Expression increased until all juxtaparanodes had immunoreactivity by P40. Developmental studies in hypomyelinating Shiverer mice revealed dramatically disrupted axoglial junctions, aberrant Na(+) channel clusters, and little or no detectable clustering of K(+) channels at all ages. These results suggest that in the optic nerve, compact myelin and normal axoglial junctions are essential for proper K(+) channel clustering and localization.

Action Potentials↗