scholarly journals Retention of glycosyltransferases in the Golgi apparatus.

1997 ◽  
Vol 44 (2) ◽  
pp. 173-179 ◽  
Author(s):  
E Jaśkiewicz
Keyword(s):  
Transmembrane Domain ◽  
Sufficient Information ◽  
Membrane Thickness ◽  
Type Ii ◽  
Transport Vesicles ◽  
Golgi Retention ◽  
Membrane Spanning ◽  
Necessary And Sufficient ◽  
Flanking Regions ◽  
Retention Signal

A number of Golgi glycosyltransferases has been cloned to date. They all are membrane proteins and share the same type II topology, but they do not possess an obvious sequence homology which would suggest a common Golgi retention signal. However, it was shown that the membrane-spanning domain and its flanking regions contain necessary and sufficient information for Golgi retention of these enzymes. Currently, two mutually complementary models have been proposed to explain the mechanism of Golgi retention of glycosyltransferases mediated by their transmembrane domain. The first model postulates the retention through oligomerization, which prevents enzymes from entering the transport vesicles. The second suggests that retention depends on the length of a membrane-spanning domain and thickness of the membrane along the Golgi complex. It has to be pointed out that neither the oligomerization nor the membrane thickness model alone can answer all questions and further work is still needed to elucidate the retention process of Golgi proteins.

scholarly journals Endoplasmic reticulum localization of Sec12p is achieved by two mechanisms: Rer1p-dependent retrieval that requires the transmembrane domain and Rer1p-independent retention that involves the cytoplasmic domain.

1996 ◽  
Vol 134 (2) ◽  
pp. 279-293 ◽  
Author(s):  
M Sato ◽  
K Sato ◽  
A Nakano
Keyword(s):  
Transmembrane Domain ◽  
Transmembrane Protein ◽  
Cytoplasmic Domain ◽  
The Other ◽  
Type Ii ◽  
Systematic Analysis ◽  
Transport Vesicles ◽  
Golgi Compartment ◽  
Retention Signal ◽  
Lines Of Evidence

Yeast Sec12p is a type II transmembrane protein in the ER, which is essential for the formation of transport vesicles. From biochemical and morphological lines of evidence, we have proposed that Sec12p is localized to the ER by two mechanisms: static retention in the ER and dynamic retrieval from the early Golgi compartment. We have also shown that Rer1p, a membrane protein in the Golgi, is required for correct localization of Sec12p. In the present study, we have performed a systematic analysis to determine the ER localization signals in Sec12p corresponding to these two mechanisms. Both the transmembrane domain (TMD) and the NH2-terminal cytoplasmic domain of Sec12p show the ability to localize the protein to the ER. The effect of the TMD is potent and sufficient by itself for the ER localization and is strongly dependent on Rer1p. On the other hand, the cytoplasmic domain shows a moderate ER-localization capability which is independent of Rer1p. The rate of mannosyl modification has been measured to distinguish between retention and retrieval. The cytoplasmic domain significantly delays the transport from the ER to the cis-Golgi. In contrast, the TMD shows only a subtle retardation in the transport from the ER to the cis-Golgi but strictly prevents the transport beyond there. From these observations, we conclude that the TMD mainly acts as the retrieval signal and the cytoplasmic domain contains the retention signal. This study not only supports the two-mechanisms hypothesis but also provides powerful tools to dissect the two.

scholarly journals The 17-residue transmembrane domain of beta-galactoside alpha 2,6-sialyltransferase is sufficient for Golgi retention

1992 ◽  
Vol 117 (2) ◽  
pp. 245-258 ◽  
Author(s):  
SH Wong ◽  
SH Low ◽  
W Hong
Keyword(s):  
Golgi Apparatus ◽  
Membrane Protein ◽  
Transmembrane Domain ◽  
Integral Membrane Protein ◽  
Chimeric Proteins ◽  
Type Ii ◽  
Lectin Affinity Chromatography ◽  
Golgi Retention ◽  
A Cell ◽  
Golgi Network

beta-Galactoside alpha 2,6-sialyltransferase (ST) is a type II integral membrane protein of the Golgi apparatus involved in the sialylation of N-linked glycans. A series of experiments has shown that the 17-residue transmembrane domain of ST is sufficient to confer localization to the Golgi apparatus when transferred to the corresponding region of a cell surface type II integral membrane protein. Lectin affinity chromatography of chimeric proteins bearing this 17-residue sequence suggests that these chimeric proteins are localized in the trans-Golgi cisternae and/or trans-Golgi network. Further experiments suggest that this 17-residue sequence functions as a retention signal for the Golgi apparatus.

scholarly journals The transmembrane domain of N-glucosaminyltransferase I contains a Golgi retention signal.

1992 ◽  
Vol 267 (14) ◽  
pp. 10122-10126 ◽  
Author(s):  
B.L. Tang ◽  
S.H. Wong ◽  
S.H. Low ◽  
W Hong
Keyword(s):  
Transmembrane Domain ◽  
Golgi Retention ◽  
Retention Signal

scholarly journals Membrane protein retention in the yeast Golgi apparatus: dipeptidyl aminopeptidase A is retained by a cytoplasmic signal containing aromatic residues.

1993 ◽  
Vol 121 (6) ◽  
pp. 1197-1209 ◽  
Author(s):  
S F Nothwehr ◽  
C J Roberts ◽  
T H Stevens
Keyword(s):  
Amino Acid ◽  
Membrane Protein ◽  
Transmembrane Domain ◽  
Mutational Analysis ◽  
Vacuolar Membrane ◽  
Type Ii ◽  
Golgi Membrane ◽  
Aromatic Residues ◽  
Golgi Retention ◽  
Dipeptidyl Aminopeptidase

The mechanism by which yeast dipeptidyl aminopeptidase (DPAP) A, type II integral membrane protein, is retained in the late Golgi apparatus has been investigated. Prior work demonstrated that the 118-amino acid cytoplasmic domain is both necessary and sufficient for Golgi retention and that mutant or overexpressed DPAP A no longer retained in the Golgi was delivered directly to the vacuolar membrane (Roberts, C. J., S. F. Nothwehr, and T. H. Stevens. 1992. J. Cell Biol. 119:69-83). Replacement of the DPAP A transmembrane domain with a synthetic hydrophobic sequence did not affect either Golgi retention of DPAP A or vacuolar delivery of the retention-defective form of DPAP A. These results indicate that the DPAP A transmembrane domain is not involved in either Golgi retention or targeting of this membrane protein. A detailed mutational analysis of the cytoplasmic domain of DPAP A indicated that the most important elements for retention were within the eight residue stretch 85-92. A 10-amino acid region from DPAP A (81-90) was sufficient for Golgi retention of alkaline phosphatase, a type II vacuolar membrane protein. Detailed mutational analysis within this 10-amino acid sufficient region demonstrated that a Phe-X-Phe-X-Asp motif was absolutely required for efficient retention. The efficiency of Golgi retention via the DPAP A signal could be diminished by overexpression of wild type but not retention-defective versions of Kex2p, another late Golgi membrane protein, suggesting that multiple Golgi membrane proteins may be retained by a common machinery. These results imply a role for a cytoplasmic signal involving aromatic residues in retention of late Golgi membrane proteins in the yeast Saccharomyces cerevisiae.

scholarly journals Localization of Sed5, a putative vesicle targeting molecule, to the cis-Golgi network involves both its transmembrane and cytoplasmic domains.

1994 ◽  
Vol 127 (2) ◽  
pp. 357-371 ◽  
Author(s):  
D K Banfield ◽  
M J Lewis ◽  
C Rabouille ◽  
G Warren ◽  
H R Pelham
Keyword(s):  
Plasma Membrane ◽  
Transmembrane Domain ◽  
Protein A ◽  
Similar Distribution ◽  
Cos Cells ◽  
Transport Vesicles ◽  
Membrane Spanning ◽  
Golgi Network ◽  
Vesicular Compartment

The yeast Sed5 protein, which is required for vesicular transport between ER and Golgi complex, is a membrane protein of the syntaxin family. These proteins are thought to provide the specific targets that are recognized by transport vesicles. We have investigated the mechanism by which Sed5 protein is itself localized. Expression of epitope-tagged versions of the yeast, Drosophila and rat Sed5 homologues in COS cells results in a perinuclear distribution; immuno-EM reveals that the majority of the protein is in a tubulo-vesicular compartment on the cis side of the Golgi apparatus. A similar distribution was obtained with a chimeric molecule consisting of a plasma membrane syntaxin with the Drosophila Sed5 transmembrane domain. This indicates that the membrane-spanning domain contains targeting information, as is the case with resident Golgi enzymes. However, alterations to the transmembrane domain of Drosophila Sed5 itself did not result in its mistargeting, implying that an additional targeting mechanism exists which involves only the cytoplasmic part of the protein. This was confirmed by modifying the transmembrane domain of the yeast Sed5 protein: substitution with the corresponding region from the Sso1 protein (a plasma membrane syntaxin homologue) did not affect yeast Sed5 function in vivo.

The role of the membrane-spanning domain and stalk region of N-acetylglucosaminyltransferase I in retention, kin recognition and structural maintenance of the Golgi apparatus in HeLa cells

1996 ◽  
Vol 109 (7) ◽  
pp. 1975-1989 ◽  
Author(s):  
T. Nilsson ◽  
C. Rabouille ◽  
N. Hui ◽  
R. Watson ◽  
G. Warren
Keyword(s):  
Golgi Apparatus ◽  
Cell Lines ◽  
Hela Cells ◽  
Kin Recognition ◽  
Dramatic Effect ◽  
Golgi Stack ◽  
Stable Cell Lines ◽  
Membrane Spanning ◽  
Necessary And Sufficient

Using a series of chimeric and truncated N-acetylglucosaminyltransferase I (NAGT I) molecules we have shown that part of the lumenal stalk region is both necessary and sufficient for kin recognition of mannosidase II and retention in the Golgi stack. The membrane-spanning domain was not required for retention, but replacing part or all of this domain with leucine residues did have a dramatic effect on Golgi morphology. In stable cell lines, stacked cisternae were replaced by tubulo-vesicular clusters containing the mutated NAGT I. The loss of stacked cisternae was proportional to the number of leucines used to replace the membrane-spanning domain.

Interaction of the endoplasmic reticulum α1,2-mannosidase Mns1p with Rer1p using the split-ubiquitin system

2001 ◽  
Vol 114 (24) ◽  
pp. 4629-4635
Author(s):  
Michel J. Massaad ◽  
Annette Herscovics
Keyword(s):  
Endoplasmic Reticulum ◽  
Catalytic Domain ◽  
Transmembrane Domain ◽  
Protein A ◽  
Yeast Cells ◽  
Type Ii ◽  
Ubiquitin System ◽  
Endoplasmic Reticulum Protein ◽  
Split Ubiquitin

The α1,2-mannosidase Mns1p involved in the N-glycosidic pathway in Saccharomyces cerevisiae is a type II membrane protein of the endoplasmic reticulum. The localization of Mns1p depends on retrieval from the Golgi through a mechanism that involves Rer1p. A chimera consisting of the transmembrane domain of Mns1p fused to the catalytic domain of the Golgi α1,2-mannosyltransferase Kre2p was localized in the endoplasmic reticulum of Δpep4 cells and in the vacuoles of rer1/Δpep4 by indirect immunofluorescence. The split-ubiquitin system was used to determine if there is an interaction between Mns1p and Rer1p in vivo. Co-expression of NubG-Mns1p and Rer1p-Cub-protein A-lexA-VP16 in L40 yeast cells resulted in cleavage of the reporter molecule, protein A-lexA-VP16, detected by western blot analysis and by expression of β-galactosidase activity. Sec12p, another endoplasmic reticulum protein that depends on Rer1p for its localization, also interacted with Rer1p using the split-ubiquitin assay, whereas the endoplasmic reticulum protein Ost1p showed no interaction. A weak interaction was observed between Alg5p and Rer1p. These results demonstrate that the transmembrane domain of Mns1p is sufficient for Rer1p-dependent endoplasmic reticulum localization and that Mns1p and Rer1p interact. Furthermore, the split-ubiquitin system demonstrates that the C-terminal of Rer1p is in the cytosol.

scholarly journals Dislocation of a Type I Membrane Protein Requires Interactions between Membrane-spanning Segments within the Lipid Bilayer

2003 ◽  
Vol 14 (9) ◽  
pp. 3690-3698 ◽  
Author(s):  
Brendan N. Lilley ◽  
Domenico Tortorella ◽  
Hidde L. Ploegh
Keyword(s):  
Membrane Protein ◽  
Heavy Chain ◽  
Transmembrane Domain ◽  
Helical Structure ◽  
Type I ◽  
Rapid Degradation ◽  
Heavy Chains ◽  
Histocompatibility Complex ◽  
Subsequent Degradation ◽  
Membrane Spanning

The human cytomegalovirus gene product US11 causes rapid degradation of class I major histocompatibility complex (MHCI) heavy chains by inducing their dislocation from the endoplasmic reticulum (ER) and subsequent degradation by the proteasome. This set of reactions resembles the endogenous cellular quality control pathway that removes misfolded or unassembled proteins from the ER. We show that the transmembrane domain (TMD) of US11 is essential for MHCI heavy chain dislocation, but dispensable for MHCI binding. A Gln residue at position 192 in the US11 TMD is crucial for the ubiquitination and degradation of MHCI heavy chains. Cells that express US11 TMD mutants allow formation of MHCI-β2m complexes, but their rate of egress from the ER is significantly impaired. Further mutagenesis data are consistent with the presence of an alpha-helical structure in the US11 TMD essential for MHCI heavy chain dislocation. The failure of US11 TMD mutants to catalyze dislocation is a unique instance in which a polar residue in the TMD of a type I membrane protein is required for that protein's function. Targeting of MHCI heavy chains for dislocation by US11 thus requires the formation of interhelical hydrogen bonds within the ER membrane.

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