Isolation and functional characterization of crustacean larval salt gland

1985 ◽  
Vol 248 (6) ◽  
pp. R702-R708
Author(s):  
R. J. Lowy ◽  
F. P. Conte
Keyword(s):  
Specific Activity ◽  
Functional Characterization ◽  
Salt Gland ◽  
Artemia Salina ◽  
The Body ◽  
Secretory Cells ◽  
Cell Integrity ◽  
Salt Glands ◽  
And Function ◽  
First Time

A batch method for isolating viable salt glands from the naupliar brine shrimp (Artemia salina) has been developed. This protocol produces a final preparation consisting of approximately 185 isolated salt glands, representing 1 X 10(4) secretory cells/g wet wt nauplii, with a final purity of 88%. Assays of cell integrity and function indicate good retention of in situ characteristics. Vital dye was excluded by 95% of the cells for at least 24 h. The O2 consumption rate was 22.7 nM O2 X min-1 X mg protein-1 and could be altered predictably by compounds known to affect oxidative phosphorylation and ion transport. The specific activity of the Na+-K+-ATPase in the salt gland, measured here for the first time, was 9.1 mM Pi X h-1 X mg protein-1. This is a substantial proportion of the body total, 17%, as expected for an active ion-transporting epithelium.

Biogenesis of Plasma Membranes in Salt Glands of Salt-Stressed Domestic Ducklings: Localization of Acyltransferase Activity

1972 ◽  
Vol 11 (3) ◽  
pp. 855-873
Author(s):  
A. M. LEVINE ◽  
JOAN A. HIGGINS ◽  
R. J. BARRNETT
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Specific Activity ◽  
Salt Gland ◽  
Secretory Cells ◽  
Salt Glands ◽  
Acyltransferase Activity ◽  
Structural Localization ◽  
Differentiated Cells ◽  
Golgi Membranes

In response to salt water stress there is a marked increase in the plasma membranes of the epithelial secretory cells of the salt glands of domestic ducklings. In the present study, the fine-structural localization of the acyltransferases involved in synthesis of phospholipids has been investigated in this tissue during this increased biogenesis of plasma membranes. The specific activity of the acyltransferases of the salt gland rose in response to salt stress, and this preceded the rapid increase in weight and cellular differentiation. After the weight increase of the gland became established, the specific activity of the acyltransferases declined, but the total activity remained constant. Salt gland tissue fixed in a mixture of glutaraldehyde and formaldehyde retained 35% of the acyltransferase activity of unfixed tissue. Cytochemical studies of the localization of acyltransferase activity in fixed and unfixed salt gland showed reaction product associated only with the lamellar membranes of the Golgi complex. This localization occurred in partially differentiated cells from salt-stressed glands to the greatest extent; and to only a small extent in cells of control tissue from unstressed salt glands. Omission of substrates resulted in absence of reaction product in association with the Golgi membranes. In addition, vesicles having limiting membranes morphologically similar to the plasma membrane occurred between the Golgi region and the plasma membrane in the partially differentiated cells. The phospholipid component of the plasma membrane appears therefore to be synthesized in association with the Golgi membranes and the membrane packaged at this site from which it moves in the form of vesicles to fuse with the pre-existing plasma membrane.

Morphology of isolated crustacean larval salt glands

1985 ◽  
Vol 248 (6) ◽  
pp. R709-R716
Author(s):  
R. J. Lowy ◽  
F. P. Conte
Keyword(s):  
Plasma Membrane ◽  
Salt Gland ◽  
Artemia Salina ◽  
Apical Plasma Membrane ◽  
Secretory Cells ◽  
Cell Contact ◽  
Salt Glands ◽  
Transmission Electron

Larval salt glands isolated from the naupliar brine shrimp (Artemia salina) were examined using light microscopy and scanning and transmission electron microscopy. These methods demonstrated that most cellular and subcellular features of the in vitro organ compared favorably with those seen in vivo. This salt gland measures 130 micron in diameter and is comprised of 50-70 secretory cells, which are of a single epithelial cell type. Characteristic ultrastructural features that are well preserved include apical to basal cell polarity, apical plasma membrane projections, and the extent of the basolateral tubular labyrinth and its association with numerous mitochondria. Some features that have been altered are a decrease in cell-cell contact, separation of septate junctions, and expansion of tubular labyrinth lumens and mitochondrial cristae. Use of this preparation has allowed examination of the salt gland cell's hemocoelic surface for the first time and provided information about the ultrastructure of the tufts formed by the apical plasma membrane.

scholarly journals Identification and functional characterization of the Piezo1 channel pore domain

2020 ◽  
pp. jbc.RA120.015905
Author(s):  
Elena D Nosyreva ◽  
David Thompson ◽  
Ruhma Syeda
Keyword(s):  
Functional Characterization ◽  
Functional Study ◽  
Heat Sensation ◽  
Physiological Processes ◽  
Structural Details ◽  
Functional Understanding ◽  
And Function ◽  
Downstream Processes ◽  
Pore Domain ◽  
First Time

Mechanotransduction is the process by which cells convert physical forces into electro-chemical responses. On a molecular scale these forces are detected by mechanically activated ion channels, which constitute the basis for hearing, touch, pain, cold and heat sensation amongst other physiological processes. Exciting high-resolution structural details of these channels are currently emerging that will eventually allow us to delineate the molecular determinants of gating and ion permeation. However, our structural-functional understanding across the family remains limited. Piezo1 is one of the largest and least understood of these channels, with various structurally identified features within its trimeric assembly. This study seeks to determine the modularity and function of Piezo1 channels by constructing deletion proteins guided by cryo EM structural knowledge. Our comprehensive functional study identified, for the first time, the minimal amino acid sequence of the full-length Piezo1 that can fold and function as the channel’s pore domain between E2172 and the last residue E2547. While, the addition of an anchor region has no effect on permeation properties. The Piezo1 pore domain is not pressure sensitive and the appending of Piezo Repeat-A did not restore pressure-dependent gating, hence the sensing module must exist between residues 1-1952. Our efforts delineating the permeation and gating regions within this complex ion channel have implications in identifying small molecules that exclusively regulate the activity of the channel’s pore module to influence mechanotransduction and downstream processes.

scholarly journals Ontogenic, phenotypic, and functional characterization of XCR1+ dendritic cells leads to a consistent classification of intestinal dendritic cells based on the expression of XCR1 and SIRPα

2014 ◽  
Author(s):  
Martina Becker ◽  
Steffen Güttler ◽  
Annabell Bachem ◽  
Evelyn Hartung ◽  
Ahmed Mora ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
T Cell ◽  
Functional Characterization ◽  
Cross Presentation ◽  
Intestinal Immune System ◽  
Lineage Marker ◽  
And Function ◽  
First Time

In the past, lack of lineage markers confounded the classification of dendritic cells (DC) in the intestine and impeded a full understanding of their location and function. We have recently shown that the chemokine receptor XCR1 is a lineage marker for cross-presenting DC in the spleen. Now we provide evidence that intestinal XCR1+ DC largely, but not fully, overlap with CD103+ CD11b- DC, the hypothesized correlate of “cross-presenting DC” in the intestine, and are selectively dependent in their development on the transcription factor Batf3. XCR1+ DC are located in the villi and epithelial crypts of the lamina propria of the small intestine, the T cell zones of Peyer’s Patches, and in the T cell zones and sinuses of the draining mesenteric lymph node. Functionally, we could demonstrate for the first time that XCR1+ / CD103+ CD11b- DC excel in the cross-presentation of orally applied antigen. Together, our data show that XCR1 is a lineage marker for cross-presenting DC also in the intestinal immune system. Further, extensive phenotypic analyses reveal that expression of the integrin SIRPα consistently demarcates the XCR1- DC population. We propose a simplified and consistent classification system for intestinal DC based on the expression of XCR1 and SIRPα.

Salt stress increases abundance and glycosylation of CFTR localized at apical surfaces of salt gland secretory cells

1994 ◽  
Vol 267 (4) ◽  
pp. C990-C1001 ◽  
Author(s):  
S. A. Ernst ◽  
K. M. Crawford ◽  
M. A. Post ◽  
J. A. Cohn
Keyword(s):  
Salt Stress ◽  
Single Cells ◽  
Salt Gland ◽  
Cell Protein ◽  
Secretory Cells ◽  
Salt Glands ◽  
Cl Secretion ◽  
Peptide Antibody ◽  
Nacl Secretion ◽  
Apical Membranes

Osmotic stress elicits hypertonic NaCl secretion and promotes structural and biochemical differentiation in avian salt glands. In addition to cholinergic control, Cl- secretion is stimulated by vasoactive intestinal peptide (VIP), suggesting that the cystic fibrosis transmembrane conductance regulator (CFTR) may be present and that its expression may be regulated by chronic salt stress. Anion efflux, assayed by 6-methoxy-N-(3-sulfopropyl)quinolinium fluorescence changes in single cells, was stimulated by VIP or 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate. Immunoblots with a COOH-terminal peptide antibody to human CFTR revealed approximately 170- and approximately 180-kDa bands in lysates from control and salt-stressed glands, respectively. Both variants reduced to approximately 140 kDa after N-glycanase digestion and gave identical tryptic phosphopeptide maps after immunoprecipitation and phosphorylation by protein kinase A. CFTR was localized to apical membranes by immunofluorescence and, additionally, to subapical vesicles by immunoelectron microscopy. Salt stress induced an approximately twofold increase in CFTR abundance/cell protein (approximately 5-fold/cell) and intensified apical membrane immunofluorescence. For comparison, Na+ pump expression increased approximately fourfold per cell protein with little change in actin. Thus differentiation induced by salt stress is accompanied by alteration in CFTR abundance and glycosylation. Upregulation of CFTR likely contributes to increased efficiency of Cl- secretion.

The Formation of the Ootheca by Periplaneta americana II. The Structure and Function of the Left Colleterial Gland

1952 ◽  
Vol s3-93 (21) ◽  
pp. 47-69
Author(s):  
P.C. J. BRUNET
Keyword(s):  
Periplaneta Americana ◽  
Secretory Activity ◽  
The Body ◽  
Secretory Cells ◽  
Structural Protein ◽  
The Right ◽  
And Function ◽  
Immature Cells ◽  
Protein Secreting ◽  
Colleterial Gland

The anatomy of the left colleterial gland of the cockroach is described. It is convenient to subdivide the gland into four regions. At the posterior end, abutting on to the outlet of the gland, there is no secretory activity and the cells of this region are not unlike normal epidermal cells. Anterior to this region are the three secretory regions of the gland; of these, the anterior and posterior secrete the structural protein and the constituent cells are equipped with a complex end-apparatus, a thick-walled depression in the apex of the cell in which the final stages of the elaboration of the secretion occur. The body of the end-apparatus contains canalicules which lead to the lumen of the gland. Alkaline phosphatase is abundantly present in this organ. Protein is secreted continuously, and there is no cycle related to oviposition. The third region, between the protein-secreting regions, secretes an oxidase, whose function may well be to oxidize the phenolic tannin-precursor produced in the right colleterial gland when the products of the two glands come together at oviposition. The gland becomes fully functional some 14 days after the final moult, the immature cells of the nymph developing directly into mature cells characteristic of the region in which they occur. Within each of the main regions the cells show some differences which suggest that there is a wave of change passing along the gland. It appears that the cells of the anterior end of the gland become defunct and the hitherto inactive cells of the posterior end take on a secretory function. The structural protein has a high phenolic content, and contains no combined carbohydrate. The presence of lipoid in the secretory cells appears to be directly bound up with the secretory processes.

scholarly journals Current Understanding of Role of Vesicular Transport in Salt Secretion by Salt Glands in Recretohalophytes

2021 ◽  
Vol 22 (4) ◽  
pp. 2203 ◽  
Author(s):  
Chaoxia Lu ◽  
Fang Yuan ◽  
Jianrong Guo ◽  
Guoliang Han ◽  
Chengfeng Wang ◽  
...  
Keyword(s):  
Vesicular Transport ◽  
Salt Gland ◽  
Soil Salinization ◽  
The Body ◽  
Molecular Evidence ◽  
Salt Glands ◽  
Functional Roles ◽  
Secretion Process ◽  
Salt Secretion

Soil salinization is a serious and growing problem around the world. Some plants, recognized as the recretohalophytes, can normally grow on saline–alkali soil without adverse effects by secreting excessive salt out of the body. The elucidation of the salt secretion process is of great significance for understanding the salt tolerance mechanism adopted by the recretohalophytes. Between the 1950s and the 1970s, three hypotheses, including the osmotic potential hypothesis, the transfer system similar to liquid flow in animals, and vesicle-mediated exocytosis, were proposed to explain the salt secretion process of plant salt glands. More recently, increasing evidence has indicated that vesicular transport plays vital roles in salt secretion of recretohalophytes. Here, we summarize recent findings, especially regarding the molecular evidence on the functional roles of vesicular trafficking in the salt secretion process of plant salt glands. A model of salt secretion in salt gland is also proposed.

scholarly journals PRESERVATION OF Na-K-ACTIVATED AND Mg-ACTIVATED ADENOSINE TRIPHOSPHATASE ACTIVITIES OF AVIAN SALT GLAND AND TELEOST GILL WITH FORMALDEHYDE AS FIXATIVE

1970 ◽  
Vol 18 (4) ◽  
pp. 251-263 ◽  
Author(s):  
STEPHEN A. ERNST ◽  
CHARLES W. PHILPOTT
Keyword(s):  
Atpase Activity ◽  
Adenosine Triphosphatase ◽  
Electron Microscopic Examination ◽  
Salt Gland ◽  
Chloride Cells ◽  
Secretory Cells ◽  
Salt Glands ◽  
Electron Microscopic ◽  
Fixed Tissue ◽  
Gill Filaments

The effect of glutaraldehyde and formaldehyde fixation on the level of biochemically demonstrable Na-K-adenosine triphosphatase (Na-K-ATPase) and Mg-ATPase of avian salt glands and teleost gill filaments was studied. Sections, 100-200 µ, prepared with the Smith-Farquhar tissue chopper, were fixed for varying periods, homogenized and assayed for ATPase activity. Fixation of salt gland tissue with 0.5% glutaraldehyde for 40-60 min completely inhibited the Na-K-ATPase activity and reduced the level of Mg-ATPase by 85%. In contrast, fixation with 2 or 3% formaldehyde, prepared from paraformaldehyde, for 60-90 min resulted in a loss of only 30% of the Na-K-ATPase activity and 65% of the Mg-ATPase activity. Similar results were obtained with gill filaments. In addition, Na-K-ATPase of formaldehyde-fixed tissue retained an obligatory requirement for Na+ and K+ and was fully sensitive to ouabain. Electron microscopic examination of formaldehyde-fixed tissue, sectioned with either the tissue chopper or in the cryostat and incubated in the Wachstein-Meisel medium, showed excellent morphologic preservation. Reaction product deposition (presumably due to Mg-ATPase) was associated with the extracellular side of the plasma membrane in the secretory cells of the salt gland and over the mitochondrial matrix of chloride cells present in the gill epithelium.

scholarly journals Transfer of human chromosomes via human minisegregant cells into mouse cells and the quantitation of the expression of hypoxanthine phosphoribosyltransferase in the hybrids

1978 ◽  
Vol 30 (1) ◽  
pp. 193-209
Author(s):  
A. Tourian ◽  
R.T. Johnson ◽  
K. Burg ◽  
S.W. Nicolson ◽  
K. Sperling
Keyword(s):  
Specific Activity ◽  
Functional Characterization ◽  
Phosphoglycerate Kinase ◽  
Kinetic Constants ◽  
Heat Inactivation ◽  
Hypoxanthine Phosphoribosyltransferase ◽  
Human Chromosomes ◽  
Intact Cells ◽  
And Function

The behaviour of human cells arrested in mitosis can be severely perturbed so as to generate numerous small minisegregants containing very few chromosomes. These cells can be separated according to size and DNA content and fused with intact cells. In this paper we describe the production and some properties of proliferating cell hybrids generated by fusion of human minisegregant cells derived from a HeLa strain with mouse A9 cells deficient in hypoxanthine phosphoribosyltransferase (HPRT, EC 2.4.2.8). The hybrids were shown to contain up to 10 human chromosomes including a single X. Independently derived hybrid clones were quantitatively characterized and compared with the parental phenotypes with respect to HPRT. Human isozymes of each of the 3 enzymes HPRT, glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and phosphoglycerate kinase (EC 2,7.2.3) were found. Tests to evaluate both structure and function of HPRT were utilized. The specific activity of HPRT of more than 10 hybrids tested was approximately 10% that of the HeLa parent. Structural characterization of HPRT from hybrid cells as evidenced by heat inactivation and electrophoretic mobility results in a ‘human-like’ phenotype. Functional characterization of parental HPRT results in kinetic constants for cofactor and substrate which do not permit distinction of human and of human and mouse enzymes; HPRT from the minisegregant hybrids had normal kinetic constants. The reduced specific activity of HPRT in the hybrids is discussed in terms of the inability of the mouse environment to regulate the full expression of the human structural gene.

scholarly journals A General Role for Rab27a in Secretory Cells

2004 ◽  
Vol 15 (1) ◽  
pp. 332-344 ◽  
Author(s):  
Tanya Tolmachova ◽  
Ross Anders ◽  
Jane Stinchcombe ◽  
Giovanna Bossi ◽  
Gillian M. Griffiths ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Knockout Mouse ◽  
Cell Types ◽  
Secretory Cells ◽  
Rab Gtpases ◽  
Specific Expression ◽  
General Role ◽  
Multistep Process ◽  
And Function ◽  
First Time

Vesicular transport is a complex multistep process regulated by distinct Rab GTPases. Here, we show for the first time that an EGFP-Rab fusion protein is fully functional in a mammalian organism. We constructed a PAC-based transgenic mouse, which expresses EGFP-Rab27a under the control of endogenous Rab27a promoter. The EGFP-Rab27a transgene was fully functional and rescued the two major defects of the ashen Rab27a knockout mouse. We achieved cell-specific expression of EGFP-Rab27a, which faithfully followed the pattern of expression of endogenous Rab27a. We found that Rab27a is expressed in an exceptionally broad range of specialized secretory cells, including exocrine (particularly in mucin- and zymogen-secreting cells), endocrine, ovarian, and hematopoietic cells, most of which undergo regulated exocytosis. We suggest that Rab27a acts in concert with Rab3 proteins in most regulated secretory events. The present strategy represents one way in which the complex pattern of expression and function of proteins involved in specialized cell types may be unraveled.

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