Localization of ral, a small Mr GTP-binding protein, to collecting duct cells in bovine and rat kidney

1991 ◽  
Vol 261 (6) ◽  
pp. F1063-F1070
Author(s):  
A. Gupta ◽  
B. Bastani ◽  
P. Chardin ◽  
K. A. Hruska
Keyword(s):  
Gene Product ◽  
Plasma Membranes ◽  
Binding Protein ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Kidney Cortex ◽  
Gtp Binding Protein ◽  
Bovine Kidney ◽  
Gtp Binding ◽  
Collecting Ducts

Plasma membranes from bovine kidney cortex were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to nitrocellulose membranes. Blotting with [alpha-32P]GTP and [35S]GTP gamma S demonstrated specific binding to three and six distinct protein bands, respectively, in the 20,000- to 29,000-Mr range. This indicated the presence of small Mr GTP binding proteins (smg) in bovine kidney cortex. Only one smg with 28,000 Mr was labeled with hydrolysis-resistant GTP photoaffinity probe p3-(4-azidoanilido)-p1-5GTP (AAGTP). The major smg in platelet membranes that binds GTP on nitrocellulose blots has been identified as ral-Mr 29,000. With the use of an antiserum against the ral A gene product, one of the smg with Mr of 29,000 present in bovine renal cortical plasma membranes was identified as ral. Ral was absent from glomerular homogenate, suggesting that it is localized to the tubular segments of the nephron. Ral was detected only in the particulate fraction and not the cytosol. Further subcellular localization of ral was investigated by immunohistochemical staining. Anti-ral antibody immunostained the apical and basolateral membranes of cells in the cortical and medullary collecting ducts in a speckled pattern in the bovine kidney. In the rat kidney, however, uniform linear staining of cortical and medullary collecting ducts predominantly localized to the apical membrane was observed. To date, no function has been assigned to ral. Localization of the ral gene product to the collecting duct suggests a specific functional role for this GTP-binding protein.

Lectin-gold cytochemistry reveals intercalated cell heterogeneity along rat kidney collecting ducts

1985 ◽  
Vol 248 (3) ◽  
pp. C348-C356 ◽  
Author(s):  
D. Brown ◽  
J. Roth ◽  
L. Orci
Keyword(s):  
Plasma Membranes ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Lectin Binding ◽  
Helix Pomatia ◽  
Intercalated Cells ◽  
Double Labeling ◽  
Thin Sections ◽  
Intercalated Cell ◽  
Collecting Ducts

The lectin-gold technique was used to detect Helix pomatia and Dolichos biflorus lectin binding sites directly on semithin and thin sections of rat kidney collecting ducts. Intercalated cell apical plasma membranes and the membranes of apical cytoplasmic vesicles were heavily labeled in the cortex and outer stripe of the outer medulla but were negative or very weakly labeled in the inner stripe and inner medulla. In contrast, clear cell apical membranes were labeled along the entire length of the collecting duct. Double labeling of semithin cryostat sections with a specific antibody and lectin-gold complexes was used to demonstrate that the intercalated cells in all regions studied contained carbonic anhydrase, even though the lectin binding differed. These results indicate that, in terms of their glycocalyx composition, intercalated cells represent a heterogeneous population in different regions of the collecting duct.

Light-induced interaction between rhodopsin and GTP-binding protein leads to the hydrolysis of GTP in the rod outer segment

1986 ◽  
Vol 64 (4) ◽  
pp. 304-308 ◽  
Author(s):  
B. D. Gupta ◽  
T. J. Borys ◽  
S. Deshpande ◽  
R. E. Jones ◽  
E. W. Abrahamson
Keyword(s):  
Light Scattering ◽  
Outer Segment ◽  
Plasma Membranes ◽  
Binding Protein ◽  
Rod Outer Segments ◽  
Particle Scattering ◽  
Gtp Binding Protein ◽  
Gtp Binding ◽  
Time Period ◽  
Hydrolysis Of

In the presence of exogeneous GTP, vertebrate whole rod outer segments (ROS), with perforated plasma membranes in the "single particle" scattering range, elicit a light-induced light-scattering transient which we call the "G" signal. Here, we report on the characteristics of the "G" signal relative to the "binding" and "dissociation" signals reported by Kuhn and colleagues. Replacing GTP with guanylyl imidodiphosphate (GMP-PNP) does not give rise to the G signal. This indicates that hydrolysis of the terminal phosphate is required for the G signal and, in addition. GTP and GMP-PNP compete for the same binding site of the enzyme responsible for the G signal (i.e., GTP-binding protein). Also, neither GDP nor its nonhydrolyzable analogue, guanosine 5′-O-(2-thiodiphosphate), when present in ROS suspensions yield any light-scattering transient in the time period tested.

Impaired chromosome partitioning and synchronization of DNA replication initiation in an insertional mutant in the Vibrio harveyi cgtA gene coding for a common GTP-binding protein

2002 ◽  
Vol 362 (3) ◽  
pp. 579-584 ◽  
Author(s):  
Monika SŁOMIṄSKA ◽  
Grażyna KONOPA ◽  
Grzegorz WĘGRZYN ◽  
Agata CZYŻ
Keyword(s):  
Cell Division ◽  
Gene Product ◽  
Vibrio Harveyi ◽  
Binding Protein ◽  
Chromosome Replication ◽  
Replication Initiation ◽  
Gtp Binding Protein ◽  
Gtp Binding ◽  
Insertional Mutant ◽  
Chromosome Partitioning

The Vibrio harveyi cgtA gene product belongs to a subfamily of small GTP-binding proteins, called Obg-like proteins. Members of this subfamily are present in diverse organisms ranging from bacteria to humans. On the other hand, the functions of these proteins in the regulation of cellular processes are largely unknown. Genes coding for these proteins are essential in almost all bacteria investigated thus far. However, a viable V. harveyi insertional mutant in the cgtA gene was described recently. Therefore, this mutant gives a unique opportunity to study functions of a member of the subfamily of Obg-like proteins. Here we demonstrate that the mutant cells often form long filaments with expanded, non-partitioned or rarely partitioned chromosomes. Such a phenotype suggests impairment of the mechanism of chromosome partition. Flow cytometric studies revealed that synchronization of chromosome replication initiation is also significantly disturbed in the cgtA mutant. Moreover, in contrast to wild-type V. harveyi, inhibition of chromosome replication and/or of cell division in the mutant bacteria caused significant increase in the number of large cells, suggesting that the cgtA gene product may be involved in the coupling of cell growth to chromosome replication and cell division. These results indicate that CgtA, an Obg-like GTP-binding protein, plays an important role in the regulation of chromosomal functions.

scholarly journals Binding of parathyroid hormone to bovine kidney-cortex plasma membranes

1973 ◽  
Vol 134 (4) ◽  
pp. 913-921 ◽  
Author(s):  
H. S. Sutcliffe ◽  
T. J. Martin ◽  
J. A. Eisman ◽  
R. Pilczyk
Keyword(s):  
Parathyroid Hormone ◽  
Adenylate Cyclase ◽  
Plasma Membranes ◽  
Specific Activity ◽  
Specific Binding ◽  
Rat Kidney ◽  
Kidney Cortex ◽  
Bovine Kidney ◽  
Hormone Sensitive ◽  
Chloramine T

1. Plasma membranes were purified from bovine kidney cortex, with a fourfold increase in specific activity of parathyroid hormone-sensitive adenylate cyclase over that in the crude homogenate. The membranes were characterized by enzyme studies. 2. Parathyroid hormone was labelled with 125I by an enzymic method and the labelled hormone shown to bind to the plasma membranes and to be specifically displaced by unlabelled hormone. Parathyroid hormone labelled by the chloramine-t procedure showed no specific binding. 75Se-labelled human parathyroid hormone, prepared in cell culture, also bound to the membranes. 3. Parathyroid hormone was shown to retain biological activity after iodination by the enzymic method, but no detectable activity remained after chloramine-t treatment. 4. High concentration of pig insulin inhibited binding of labelled parathyroid hormone to plasma membranes and partially inhibited the hormone-sensitive adenylate cyclase activity in a crude kidney-cortex preparation. 5. EDTA enhanced and Ca2+ inhibited binding of labelled parathyroid hormone to plasma membranes. 6. Whereas rat kidney homogenates were capable of degrading labelled parathyroid hormone to trichloroacetic acid-soluble fragments, neither crude homogenates nor purified membranes from bovine kidney showed this property. 7. Binding of parathyroid hormone is discussed in relation to metabolism and initial events in hormone action.

scholarly journals Purification and subunit composition of a GTP-binding protein from maize root plasma membranes

FEBS Letters ◽  
1991 ◽  
Vol 291 (2) ◽  
pp. 219-221 ◽  
Author(s):  
Sevo V. Bilushi ◽  
Alexander G. Shebunin ◽  
Alexey V. Babakov
Keyword(s):  
Plasma Membranes ◽  
Binding Protein ◽  
Maize Root ◽  
Subunit Composition ◽  
Gtp Binding Protein ◽  
Gtp Binding

The small GTP-binding protein, Rab6p, is associated with both Golgi and post-Golgi synaptophysin-containing membranes during synaptogenesis of hypothalamic neurons in culture

1993 ◽  
Vol 105 (4) ◽  
pp. 935-947 ◽  
Author(s):  
A. Tixier-Vidal ◽  
A. Barret ◽  
R. Picart ◽  
V. Mayau ◽  
D. Vogt ◽  
...  
Keyword(s):  
Synaptic Vesicles ◽  
Laser Scanning ◽  
Synapse Formation ◽  
Binding Protein ◽  
Rat Kidney ◽  
Integral Membrane Protein ◽  
Laser Scanning Confocal Microscopy ◽  
Gtp Binding Protein ◽  
Scanning Confocal Microscopy ◽  
Gtp Binding

We have recently localized a small GTP-binding protein (Rab6p) thought to be involved in vesicular membrane transport, to the medial and trans-cisternae of the Golgi apparatus in NRK (normal rat kidney) cells. Here, we have localized and quantified Rab6p during the development in culture of embryonic neurons, up to synapse formation, and compared its subcellular distribution and level of expression to that of synaptophysin, a major integral membrane protein of small synaptic vesicles. Using immunocytochemistry (laser scanning confocal microscopy, immunoelectron microscopy), fractionation and immunoisolation methods, we show that during the early phase of synaptogenesis, Rab6p is associated with synaptophysin-containing membranes of a trans-Golgi subcompartment, post-Golgi vesicles and small synaptic vesicles or their precursors. Concomitantly, Rab6p undergoes translocation from cytosol to membranes and its level of expression increases. However, at late stages, the association of Rab6p to small synaptic vesicles sharply decreases and its level of expression plateaus. These findings suggest a role for Rab6p in the post-Golgi transport of synaptophysin, at an early step of the biogenesis of small synaptic vesicles.

scholarly journals A blue-light-activated GTP-binding protein in the plasma membranes of etiolated peas.

1991 ◽  
Vol 88 (20) ◽  
pp. 8925-8929 ◽  
Author(s):  
K. M. Warpeha ◽  
H. E. Hamm ◽  
M. M. Rasenick ◽  
L. S. Kaufman
Keyword(s):  
Blue Light ◽  
Plasma Membranes ◽  
Binding Protein ◽  
Gtp Binding Protein ◽  
Gtp Binding
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