NBD-labeled phosphatidylcholine enters the yeast vacuole via the pre-vacuolar compartment

2002 ◽  
Vol 115 (13) ◽  
pp. 2725-2733 ◽  
Author(s):  
Pamela K. Hanson ◽  
Althea M. Grant ◽  
J. Wylie Nichols
Keyword(s):  
Plasma Membrane ◽  
Low Temperature ◽  
Head Group ◽  
Temperature Sensitive ◽  
Vesicular Traffic ◽  
Yeast Vacuole ◽  
Golgi Transport ◽  
Vacuolar Compartment ◽  
Wild Type Yeast ◽  
Similar Enrichment

At low temperature, the short-chain fluorescent-labeled phospholipids,1-myristoyl-2-[6-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)aminocaproyl]-phosphatidylcholine (M-C6-NBD-PC) and its phosphatidylethanolamine analog, M-C6-NBD-PE, are internalized by flip across the plasma membrane of S. cerevisiae and show similar enrichment in intracellular membranes including the mitochondria and nuclear envelope/ER. At higher temperatures (24-37°C), or if low temperature internalization is followed by warming, M-C6-NBD-PC, but not M-C6-NBD-PE, is trafficked to the lumen of the vacuole. Sorting of M-C6-NBD-PC to the vacuole is blocked by energy-depletion and by null mutations in the VPS4 and VPS28 genes required for vesicular traffic from the pre-vacuolar compartment (PVC) to the vacuole. This sorting is not blocked by a temperature-sensitive mutation in SEC12,which inhibits ER to Golgi transport, a null mutation in VPS8, which inhibits Golgi to PVC transport, or temperature-sensitive and null mutations in END4, which inhibit endocytosis from the plasma membrane. Monomethylation or dimethylation of the primary amine head-group of M-C6-NBD-PE is sufficient for sorting to the yeast vacuole in both wild-type yeast and in strains defective in the phosphatidylethanolamine methylation pathway. These data indicate that methylation of M-C6-NBD-PE produces the crucial structural component required to sort these phospholipid analogues to the vacuole via the PVC.

3-O-methyl-D-glucose transport in tumoral insulin-producing cells

1986 ◽  
Vol 251 (6) ◽  
pp. C841-C846 ◽  
Author(s):  
W. J. Malaisse ◽  
M. H. Giroix ◽  
F. Malaisse-Lagae ◽  
A. Sener
Keyword(s):  
Plasma Membrane ◽  
Low Temperature ◽  
Glucose Transport ◽  
Concentration Gradient ◽  
Glucose Oxidation ◽  
Temperature Sensitive ◽  
Insulin Producing Cells ◽  
Different Temperatures ◽  
High Concentrations ◽  
Tumoral Cells

Tumoral insulin-producing cells of the RINm5F line were exposed at different temperatures, and for various lengths of time to increasing concentrations of 3-O-methyl-D-[U-14C]glucose. The uptake of the hexose represented a temperature-sensitive and saturable process, so that no rapid equilibration of hexose concentrations across the plasma membrane was reached, especially at low temperature and/or high concentrations of 3-O-methyl-D-glucose. The uptake of 3-O-methyl-D-[U-14C]glucose was not affected by a prior loading of the cells with the unlabeled hexose and its release from prelabeled cells was observed in the absence of any concentration gradient across the plasma membrane. The uptake of D-[U-14C]glucose and utilization of D-[5-3H]glucose was inhibited by 3-O-methyl-D-glucose, which failed, however, to affect D-[U-14C]glucose oxidation. At variance with the situation found in normal insulin-producing cells, the transport of D-glucose into the tumoral cells may thus play a regulatory role in its metabolism.

scholarly journals High-Copy Suppressor Analysis Reveals a Physical Interaction between Sec34p and Sec35p, a Protein Implicated in Vesicle Docking

1999 ◽  
Vol 10 (10) ◽  
pp. 3317-3329 ◽  
Author(s):  
Dong-Wook Kim ◽  
Michael Sacher ◽  
Al Scarpa ◽  
Anne Marie Quinn ◽  
Susan Ferro-Novick
Keyword(s):  
Temperature Sensitive ◽  
Physical Interaction ◽  
Synthetic Lethal ◽  
Vesicle Docking ◽  
Vesicular Traffic ◽  
Golgi Transport ◽  
Synthetic Lethal Interactions ◽  
Golgi Protein ◽  
Late Stages ◽  
Suppressor Analysis

A temperature-sensitive mutant, sec34-2, is defective in the late stages of endoplasmic reticulum (ER)-to-Golgi transport. A high-copy suppressor screen that uses thesec34-2 mutant has resulted in the identification of theSEC34 structural gene and a novel gene calledGRP1. GRP1 encodes a previously unidentified hydrophilic yeast protein related to the mammalian Golgi protein golgin-160. Although GRP1 is not essential for growth, the grp1Δ mutation displays synthetic lethal interactions with several mutations that result in ER accumulation and a block in the late stages of ER-to-Golgi transport, but not with those that block the budding of vesicles from the ER. Our findings suggest that Grp1p may facilitate membrane traffic indirectly, possibly by maintaining Golgi function. In an effort to identify genes whose products physically interact with Sec34p, we also tested the ability of overexpressed SEC34 to suppress known secretory mutations that block vesicular traffic between the ER and the Golgi. This screen revealed that SEC34 specifically suppressessec35-1. SEC34 encodes a hydrophilic protein of ∼100 kDa. Like Sec35p, which has been implicated in the tethering of ER-derived vesicles to the Golgi, Sec34p is predominantly soluble. Sec34p and Sec35p stably associate with each other to form a multiprotein complex of ∼480 kDa. These data indicate that Sec34p acts in conjunction with Sec35p to mediate a common step in vesicular traffic.

scholarly journals Efforts towards a low-temperature-sensitive physics package for vapor cell atomic clocks

2020 ◽  
Vol 1 (1) ◽  
Author(s):  
Qiang Hao ◽  
Wenxiang Xue ◽  
Feng Xu ◽  
Kemu Wang ◽  
Peter Yun ◽  
...  
Keyword(s):  
Low Temperature ◽  
Temperature Sensitive ◽  
Atomic Clocks ◽  
Vapor Cell

Development of low temperature curing, 120°C, durable, corrosion protection powder coatings for temperature sensitive substrates

2005 ◽  
Vol 2 (8) ◽  
pp. 661-668 ◽  
Author(s):  
Glen Merfeld ◽  
Steve Mordhorst ◽  
Rainer Koeniger ◽  
A. Ersin Acar ◽  
Chris Molaison ◽  
...  
Keyword(s):  
Low Temperature ◽  
Corrosion Protection ◽  
Temperature Sensitive ◽  
Powder Coatings

scholarly journals Vesicular Transport Is Not Required for the Cytoplasmic Pool of Cholera Toxin To Interact with the Stimulatory Alpha Subunit of the Heterotrimeric G Protein

2004 ◽  
Vol 72 (12) ◽  
pp. 6826-6835 ◽  
Author(s):  
Ken Teter ◽  
Michael G. Jobling ◽  
Randall K. Holmes
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Cholera Toxin ◽  
G Protein ◽  
Vesicular Transport ◽  
Cytotoxic Action ◽  
Anterograde Transport ◽  
Heterotrimeric G Protein ◽  
Α Subunit ◽  
Vesicular Traffic

ABSTRACT Cholera toxin (CT) moves from the cell surface to the endoplasmic reticulum (ER) by retrograde vesicular transport. The catalytic A1 polypeptide of CT (CTA1) then crosses the ER membrane, enters the cytosol, ADP-ribosylates the stimulatory α subunit of the heterotrimeric G protein (Gsα) at the cytoplasmic face of the plasma membrane, and activates adenylate cyclase. The cytosolic pool of CTA1 may reach the plasma membrane and its Gsα target by traveling on anterograde-directed transport vesicles. We examined this possibility with the use of a plasmid-based transfection system that directed newly synthesized CTA1 to either the ER lumen or the cytosol of CHO cells. Such a system allowed us to bypass the CT retrograde trafficking itinerary from the cell surface to the ER. Previous work has shown that the ER-localized pool of CTA1 is rapidly exported from the ER to the cytosol. Expression of CTA1 in either the ER or the cytosol led to the activation of Gsα, and Gsα activation was not inhibited in transfected cells exposed to drugs that inhibit vesicular traffic. Thus, anterograde transport from the ER to the plasma membrane is not required for the cytotoxic action of CTA1.

scholarly journals A High Copy Suppressor Screen Reveals Genetic Interactions Between BET3 and a New Gene: Evidence for a Novel Complex in ER-to-Golgi Transport

Genetics ◽  
1998 ◽  
Vol 149 (2) ◽  
pp. 833-841
Author(s):  
Yu Jiang ◽  
Al Scarpa ◽  
Li Zhang ◽  
Shelly Stone ◽  
Ed Feliciano ◽  
...  
Keyword(s):  
Growth Defect ◽  
Carboxypeptidase Y ◽  
Temperature Sensitive ◽  
Yeast Saccharomyces Cerevisiae ◽  
Specific Suppression ◽  
Golgi Transport ◽  
New Gene ◽  
Insight Into ◽  
Suppressor Screen

Abstract The BET3 gene in the yeast Saccharomyces cerevisiae encodes a 22-kD hydrophilic protein that is required for vesicular transport between the ER and Golgi complex. To gain insight into the role of Bet3p, we screened for genes that suppress the growth defect of the temperature-sensitive bet3 mutant at 34°. This high copy suppressor screen resulted in the isolation of a new gene, called BET5. BET5 encodes an essential 18-kD hydrophilic protein that in high copy allows growth of the bet3-1 mutant, but not other ER accumulating mutants. This strong and specific suppression is consistent with the fact that Bet3p and Bet5p are members of the same complex. Using PCR mutagenesis, we generated a temperature-sensitive mutation in BET5 (bet5-1) that blocks the transport of carboxypeptidase Y to the vacuole and prevents secretion of the yeast pheromone α-factor at 37°. The precursor forms of these proteins that accumulate in this mutant are indicative of a block in membrane traffic between the ER and Golgi apparatus. High copy suppressors of the bet5-1 mutant include several genes whose products are required for ER-to-Golgi transport (BET1, SEC22, USO1 and DSS4) and the maintenance of the Golgi (ANP1). These findings support the hypothesis that Bet5p acts in conjunction with Bet3p to mediate a late stage in ER-to-Golgi transport. The identification of mammalian homologues of Bet3p and Bet5p implies that the Bet3p/Bet5p complex is highly conserved in evolution.

EGF-and NGF-stimulated translocation of cytohesin-1 to the plasma membrane of PC12 cells requires PI 3-kinase activation and a functional cytohesin-1 PH domain

1999 ◽  
Vol 112 (12) ◽  
pp. 1957-1965 ◽  
Author(s):  
K. Venkateswarlu ◽  
F. Gunn-Moore ◽  
J.M. Tavare ◽  
P.J. Cullen
Keyword(s):  
Plasma Membrane ◽  
Pc12 Cells ◽  
Laser Scanning ◽  
Time Course ◽  
Fluorescent Protein ◽  
Dominant Negative ◽  
Head Group ◽  
Nucleotide Exchange ◽  
Ph Domain

ADP-ribosylation factors (ARFs) are small GTP-binding proteins that function as regulators of eukaryotic vesicle trafficking. Cytohesin-1 is a member of a family of ARF guanine nucleotide-exchange factors that contain a C-terminal pleckstrin homology (PH) domain which has been proposed to bind the lipid second messenger phosphatidylinositol 3,4,5-trisphosphate (PIP3). Here we demonstrate that in vitro, recombinant cytohesin-1 binds, via its PH domain, the inositol head group of PIP3, inositol 1,3,4, 5-tetrakisphosphate (IP4), with an affinity greater than 200-fold higher than the inositol head group of either phosphatidylinositol 4, 5-bisphosphate or phosphatidylinositol 3,4-bisphosphate. Moreover, addition of glycerol or diacetylglycerol to the 1-phosphate of IP4 does not alter the ability to interact with cytohesin-1, data which is entirely consistent with cytohesin-1 functioning as a putative PIP3 receptor. To address whether cytohesin-1 binds PIP3 in vivo, we have expressed a chimera of green fluorescent protein (GFP) fused to the N terminus of cytohesin-1 in PC12 cells. Using laser scanning confocal microscopy we demonstrate that either EGF- or NGF-stimulation of transiently transfected PC12 cells results in a rapid translocation of GFP-cytohesin-1 from the cytosol to the plasma membrane. This translocation is dependent on the cytohesin-1 PH domain and occurs with a time course that parallels the rate of plasma membrane PIP3 production. Furthermore, the translocation requires the ability of either agonist to activate PI 3-kinase, since it is inhibited by wortmannin (100 nM), LY294002 (50 microM) and by coexpression with a dominant negative p85. This data therefore suggests that in vivo cytohesin-1 can interact with PIP3 via its PH domain.

Cytogenetic Analyses of PSL1 Mutant, a Novel Low‐Temperature‐Sensitive Purple‐Striped Leaf Color Mutant in Wheat

Crop Science ◽  
2018 ◽  
Vol 58 (5) ◽  
pp. 1919-1931 ◽  
Author(s):  
Cong Liu ◽  
Narong Shi ◽  
Huiyu Wu ◽  
Xuyao An ◽  
Jinjuan Zheng ◽  
...  
Keyword(s):  
Low Temperature ◽  
Temperature Sensitive ◽  
Leaf Color ◽  
Color Mutant ◽  
Cytogenetic Analyses ◽  
Leaf Color Mutant
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