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1.
Calcium fluxes across the sarcoplasmic reticulum membrane are regulated by phosphorylation of a 27,000-dalton membrane-bound protein termed phospholamban. Phospholamban is phosphorylated by three different protein kinases (cAMP-dependent, Ca2+.CAM-dependent and Ca2+.phospholipid dependent) at apparently distinct sites. Phosphorylation by each of the protein kinases increases the rates of active calcium transport by sarcoplasmic reticulum vesicles. The stimulatory effects of protein kinases on the calcium pump may be reversed by an endogenous protein phosphatase activity. The phosphoprotein phosphatase can dephosphorylate both the cAMP-dependent and the Ca2+.CAM-dependent sites of phospholamban. Phosphorylation of phospholamban also occurs in situ, in perfused beating hearts, during the peak of the inotropic response to beta-adrenergic stimulation. Reversal of the stimulatory effects is associated with dephosphorylation of phospholamban. Thus, in vivo and in vitro studies suggest that phospholamban is a regulator for the calcium pump in cardiac sarcoplasmic reticulum. The degree of phospholamban phosphorylation determined by the interaction of specific protein kinases and phosphatases may represent an important control for sarcoplasmic reticulum function and, thus, for the contraction-relaxation cycle in the myocardium. In this review, we summarize recent evidence on physical and structural properties of phospholamban, the proposed structural molecular models for this protein, and the significance of its regulatory role both in vitro and in situ.  相似文献   
2.
Summary Canine cardiac sarcoplasmic reticulum is phosphorylated by adenosine 3,5-monophosphate (cAMP)-dependent and by calcium · calmodulin-dependent protein kinases on a 27 000 proteolipid, called phospholamban. Both types of phosphorylation are associated with an increase in the initial rates of Ca2+ transport by SR vesicles which reflects an increased turnover of elementary steps of the calcium ATPase reaction sequence. The stimulatory effects of the protein kinases on the calcium pump may be reversed by an endogenous protein phosphatase, which can dephosphorylate both the CAMP-dependent and the calcium · calmodulin-dependent sites on phospholamban. Thus, the calcium pump in cardiac sarcoplasmic reticulum appears to be under reversible regulation mediated by protein kinases and protein phosphatases.  相似文献   
3.
Phospholamban, the putative regulator of the Ca2+-ATPase in cardiac sarcoplasmic reticulum, was immunolocalized in canine visceral and vascular smooth muscle. Gently disrupted tissues were labeled with an affinity-purified phospholamban polyclonal antibody and indirect immunogold, using preembedding techniques. The sarcoplasmic reticulum of smooth muscle cells was specifically labeled with patches of immunogold distributed in a nonuniform fashion, while the sarcolemma did not appear to contain any phospholamban. The outer nuclear envelopes were also observed to be heavily labeled with the affinity-purified phospholamban polyclonal antibody. These findings suggest that phospholamban may play a role in the regulation of cytoplasmic and intranuclear calcium levels in smooth muscle cells.  相似文献   
4.
We present a cladistic analysis of the Cirripedia Thoracica using morphological characters and the Acrothoracica and Ascothoracida as outgroups. The list of characters comprised 32 shell and soft body features. The operational taxonomic units (OTUs) comprised 26 well-studied fossil and extant taxa, principally genera, since uncertainty about monophyly exists for most higher ranking taxonomic units. Parsimony analyses using PAUP 3.1.1 and Hennig86 produced 189 trees of assured minimal length. We also examined character evolution in the consensus trees using MacClade and Clados. The monophyly of the Balanomorpha and the Verrucomorpha sensu stricto is confirmed, and all trees featured a sister group relationship between the ‘living fossil Neoverruca and me Brachylepadomorpha. In the consensus trees the sequential progression of ‘pedunculate‘sister groups up to a node containing Neolepas also conforms to current views, but certain well-established taxa based solely on plesiomorphies stand out as paraphyletic, such as Pedunculata (= Lepadomorpha); Eolepadinae, Scalpellomorpha and Chthamaloidea. The 189 trees differed principally in the position of shell-less pedunculates, Neoverruca, the scalpelloid Capitulum, and the interrelationships within the Balanomorpha, although the 50% majority rule consensus tree almost fully resolved the latter. A monophyletic Sessilia comprising both Verrucomorpha and Balanomorpha appeared among the shortest trees, but not in the consensus. A tree with a monophyletic Verrucomorpha including Neoverruca had a tree length two steps longer than the consensus trees. Deletion of all extinct OTUs produced a radically different tree, which highlights the importance of fossils in estimating cirripede phylogeny. Mapping of our character set onto a manually constructed cladogram reflecting die most recent scenario of cirripede evolution resulted in a tree length five steps longer than any of our shortest trees. Our analysis reveals that several key questions in cirripede phylogeny remain unsolved, notably the position of shell-less forms and the transition from ‘pedunculate‘to ‘sessile‘barnacles. The inclusion of more fossil species at this point in our understanding of cirripede phylogeny will only result in even greater levels of uncertainty. When constructing the character list we also identified numerous uncertainties in the homology of traits commonly used in discussing cirripede evolution. Our study highlights larval ultrastructure, detailed studies of early ontogeny, and molecular data as the most promising areas for future research.  相似文献   
5.
The Ca2(+)-ATPase in cardiac sarcoplasmic reticulum (SR) is under regulation by phospholamban, an oligomeric proteolipid. To determine the molecular mechanism by which phospholamban regulates the Ca2(+)-ATPase, a reconstitution system was developed, using a freeze-thaw sonication procedure. The best rates of Ca2+ uptake (700 nmol/min/mg reconstituted vesicles compared with 800 nmol/min/mg SR vesicles) were observed when cholate and phosphatidylcholine were used at a ratio of cholate/phosphatidylcholine/Ca2(+)-ATPase of 2:80:1. The EC50 values for Ca2+ were 0.05 microM for both Ca2+ uptake and Ca2(+)-ATPase activity in the reconstituted vesicles compared with 0.63 microM Ca2+ in native SR vesicles. Inclusion of phospholamban in the reconstituted vesicles was associated with a significant inhibition of the initial rates of Ca2+ uptake at pCa 6.0. However, phosphorylation of phospholamban by the catalytic subunit of the cAMP-dependent protein kinase reversed the inhibitory effect on the Ca2+ pump. Similar findings were observed when a peptide, corresponding to amino acids 1-25 of phospholamban, was used. These findings indicate that phospholamban is an inhibitor of the Ca2(+)-ATPase in cardiac SR and phosphorylation of phospholamban relieves this inhibition. The mechanism by which phospholamban inhibits the Ca2+ pump is unknown, but our findings with the synthetic peptide suggest that a direct interaction between the Ca2(+)-ATPase and the hydrophilic portion of phospholamban may be one of the mechanisms for regulation.  相似文献   
6.
Both fibronectin and laminin were found by immunofluorescence as a matrix at the surface of normal rat kidney cells. These matrices were absent from the surface of virally transformed rat kidney cells. Soluble fibronectin and laminin were detected in the culture media of the transformed as well as the normal cells. Culture supernates of the transformed cells contained even more fibronectin than the supernates of the transformed cells contained even more fibronectin than the supernates of the normal cells while laminin was present in similar amounts in both culture media. This shows that the loss of fibronectin and laminin from the surface of the transformed cells is caused by failure of the cells to deposit these proteins into an insoluble matrix and not caused by inadequate production. Fibronectins isolated from culture media of the normal and transformed cells were similar in SDS polyacrylamide gel electrophresis. Laminin isolated from culture media by affinity chromatography on heparin-Sepharose followed by immunoprecipitation was composed of three main polypeptides, one with a molecular weight of 400,000 and two with a molecular weight close to 200,000 in both cell types. Fibronectins from both cell types were equally active in promoting cell attachment. Rat fibronectin from transformed cells, like normal cells, when applied to culture dishes coated with fibronectin, readily attached and spread on the substratum, requiring approximately the same amount of fibronectin as the normal cells. On the basis of these results it seem that the failure of the transformed cells to incorporate fibronectin into an insoluble cell surface matix is not a consequence of a demonstrable change in the functional characteristics of the fibronectin molecule or in the ability of the cells to interact with fibronectin. It may depend on as yet unidentified interactions of the cell surface. Similar interactions may be needed for the deposition of laminin into the matrix, because laminin was also absent from the surface of transformed cells, despite its being synthesized by these cells.  相似文献   
7.
The relationship between intracellular lysosomal rupture and cell death caused by silica was studied in P388d(1) macrophages. After 3 h of exposure to 150 μg silica in medium containing 1.8 mM Ca(2+), 60 percent of the cells were unable to exclude trypan blue. In the absence of extracellular Ca(2+), however, all of the cells remained viable. Phagocytosis of silica particles occurred to the same extent in the presence or absence of Ca(2+). The percentage of P388D(1) cells killed by silica depended on the dose and the concentration of Ca(2+) in the medium. Intracellular lyosomal rupture after exposure to silica was measured by acridine orange fluorescence or histochemical assay of horseradish peroxidase. With either assay, 60 percent of the cells exposed to 150 μg silica for 3 h in the presence of Ca(2+) showed intracellular lysosomal rupture, was not associated with measureable degradation of total DNA, RNA, protein, or phospholipids or accelerated turnover of exogenous horseradish peroxidase. Pretreatment with promethazine (20 μg/ml) protected 80 percent of P388D(1) macrophages against silica toxicity although lysosomal rupture occurred in 60-70 percent of the cells. Intracellular lysosomal rupture was prevented in 80 percent of the cells by pretreatment with indomethacin (5 x 10(-5)M), yet 40-50 percent of the cells died after 3 h of exposure to 150 μg silica in 1.8 mM extracellular Ca(2+). The calcium ionophore A23187 also caused intracellular lysosomal rupture in 90-98 percent of the cells treated for 1 h in either the presence or absence of extracellular Ca(2+). With the addition of 1.8 mM Ca(2+), 80 percent of the cells was killed after 3 h, whereas all of the cells remained viable in the absence of Ca(2+). These experiments suggest that intracellular lysosomal rupture is not causally related to the cell death cause by silica or A23187. Cell death is dependent on extracellular Ca(2+) and may be mediated by an influx of these ions across the plasma membrane permeability barrier damaged directly by exposure to these toxins.  相似文献   
8.
Summary

Cirripedes are fascinating models for studying both functional constraints and diversity in larval development. Adult cirripedes display an amazing variation in morphology from sessile suspension feeders that still retain many crustacean characters to parasites that have lost virtually all arthropod traits. In contrast, cirripede larval development follows a common scheme with pelagic larvae comprising a series of nauplii followed by a cyprid. Variations are mostly concerned with whether or not the nauplii are feeding and the degree of abbreviation of development, culminating in species where the larvae hatch as cyprids. The cypris larvae are very similar among the ingroups of the Cirripedia, but interesting variations occur in structures used for substrate location and attachment. The cyprid is specialized to both swim through the water and actively explore the substratum by walking on the antennules and using an array of sensory organs in search for a suitable site to attach. This unique morphology and behavior of the cyprid have enabled the Cirripedia to colonize widely different habitats ranging from hard rock to soft animal tissue. Yet, the cyprid can metamorphose into juveniles as different as a setose feeding barnacle and the vermiform stages of the parasitic forms. This emphasizes the importance of the cyprid as one of the key features for the evolutionary success of the Cirripedia.  相似文献   
9.
Rate control analysis defines the in vivo control map governing yeast protein synthesis and generates an extensively parameterized digital model of the translation pathway. Among other non‐intuitive outcomes, translation demonstrates a high degree of functional modularity and comprises a non‐stoichiometric combination of proteins manifesting functional convergence on a shared maximal translation rate. In exponentially growing cells, polypeptide elongation (eEF1A, eEF2, and eEF3) exerts the strongest control. The two other strong control points are recruitment of mRNA and tRNAi to the 40S ribosomal subunit (eIF4F and eIF2) and termination (eRF1; Dbp5). In contrast, factors that are found to promote mRNA scanning efficiency on a longer than‐average 5′untranslated region (eIF1, eIF1A, Ded1, eIF2B, eIF3, and eIF5) exceed the levels required for maximal control. This is expected to allow the cell to minimize scanning transition times, particularly for longer 5′UTRs. The analysis reveals these and other collective adaptations of control shared across the factors, as well as features that reflect functional modularity and system robustness. Remarkably, gene duplication is implicated in the fine control of cellular protein synthesis.  相似文献   
10.
To determine whether selective impairment of cardiac sarcoplasmic reticulum (SR) Ca(2+) transport may drive the progressive functional deterioration leading to heart failure, transgenic mice, overexpressing a phospholamban Val(49) --> Gly mutant (2-fold), which is a superinhibitor of SR Ca(2+)-ATPase affinity for Ca(2+), were generated, and their cardiac phenotype was examined longitudinally. At 3 months of age, the increased EC(50) level of SR Ca(2+) uptake for Ca(2+) (0.67 +/- 0.09 microm) resulted in significantly higher depression of cardiomyocyte rates of shortening (57%), relengthening (31%), and prolongation of the Ca(2+) signal decay time (165%) than overexpression (2-fold) of wild type phospholamban (68%, 64%, and 125%, respectively), compared with controls (100%). Echocardiography also revealed significantly depressed function and impaired beta-adrenergic responses in mutant hearts. The depressed contractile parameters were associated with left ventricular remodeling, recapitulation of fetal gene expression, and hypertrophy, which progressed to dilated cardiomyopathy with interstitial tissue fibrosis and death by 6 months in males. Females also had ventricular hypertrophy at 3 months but exhibited normal systolic function up to 12 months of age. These results suggest a causal relationship between defective SR Ca(2+) cycling and cardiac remodeling leading to heart failure, with a gender-dependent influence on the time course of these alterations.  相似文献   
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