Investigation of the requirements for transitional endoplasmic reticulum (tER) site structure and function in Saccharomyces cerevisiae.

Secretory proteins are exported from the endoplasmic reticulum (ER) at specialized regions known as the transitional ER (tER). Coat protein complex II (COPII) proteins are enriched at tER sites, although the mechanisms underlying tER site assembly and maintenance are not understood. Here, we investigated the dynamic properties of tER sites in Saccharomyces cerevisiae and probed protein and lipid requirements for tER site structure and function. Thermosensitive sec12 and sec16 mutations caused a collapse of tER sites in a manner that depended on nascent secretory cargo. Continual fatty acid synthesis was required for ER export and for normal tER site structure, whereas inhibition of sterol and ceramide synthesis produced minor effects. An in vitro assay to monitor assembly of Sec23p-green fluorescent protein at tER sites was established to directly test requirements. tER sites remained active for ∼10 min in vitro and depended on Sec12p function. Bulk phospholipids were also required for tER site structure and function in vitro, whereas depletion of phophatidylinositol selectively inhibited coat protein complex II (COPII) budding but not assembly of tER site structures. These results indicate that tER sites persist through relatively stringent treatments in which COPII budding was strongly inhibited. We propose that tER site structures are stable elements that are assembled on an underlying protein and lipid scaffold.

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The Schizosaccharomyces pombe cho1+ Gene Encodes a Phospholipid Methyltransferase

Genetics ◽

10.1093/genetics/150.2.553 ◽

1998 ◽

Vol 150 (2) ◽

pp. 553-562

Author(s):

Margaret I Kanipes ◽

John E Hill ◽

Susan A Henry

Keyword(s):

Saccharomyces Cerevisiae ◽

Sequence Analysis ◽

Schizosaccharomyces Pombe ◽

Gene Disruption ◽

Structure And Function ◽

Biosynthetic Enzyme ◽

Gene Encoding ◽

Complementation Groups ◽

Eukaryotic Organisms ◽

And Function

Abstract The isolation of mutants of Schizosaccharomyces pombe defective in the synthesis of phosphatidylcholine via the methylation of phosphatidylethanolamine is reported. These mutants are choline auxotrophs and fall into two unlinked complementation groups, cho1 and cho2. We also report the analysis of the cho1+ gene, the first structural gene encoding a phospholipid biosynthetic enzyme from S. pombe to be cloned and characterized. The cho1+ gene disruption mutant (cho1Δ) is viable if choline is supplied and resembles the cho1 mutants isolated after mutagenesis. Sequence analysis of the cho1+ gene indicates that it encodes a protein closely related to phospholipid methyltransferases from Saccharomyces cerevisiae and rat. Phospholipid methyltransferases encoded by a rat liver cDNA and the S. cerevisiae OPI3 gene are both able to complement the choline auxotrophy of the S. pombe cho1 mutants. These results suggest that both the structure and function of the phospholipid N-methyltransferases are broadly conserved among eukaryotic organisms.

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Structure and function of IP3 receptor, Ca2+ channel on the endoplasmic reticulum

Seibutsu Butsuri ◽

10.2142/biophys.43.s8_2 ◽

2003 ◽

Vol 43 (supplement) ◽

pp. S8

Author(s):

K. Mikoshiba

Keyword(s):

Endoplasmic Reticulum ◽

Structure And Function ◽

Ip3 Receptor ◽

And Function

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Oxidative protein folding in the mammalian endoplasmic reticulum

Biochemical Society Transactions ◽

10.1042/bst0320655 ◽

2004 ◽

Vol 32 (5) ◽

pp. 655-658 ◽

Cited By ~ 51

Author(s):

C.E. Jessop ◽

S. Chakravarthi ◽

R.H. Watkins ◽

N.J. Bulleid

Keyword(s):

Endoplasmic Reticulum ◽

Mammalian Cells ◽

Redox State ◽

Structure And Function ◽

Reduced Form ◽

Secretory Proteins ◽

Oxidative Protein Folding ◽

Disulphide Bonds ◽

And Function ◽

Substrate Proteins

Native disulphide bonds are essential for the structure and function of many membrane and secretory proteins. Disulphide bonds are formed, reduced and isomerized in the endoplasmic reticulum of mammalian cells by a family of oxidoreductases, which includes protein disulphide isomerase (PDI), ERp57, ERp72, P5 and PDIR. This review will discuss how these enzymes are maintained in either an oxidized redox state that allows them to form disulphide bonds in substrate proteins or a reduced form that allows them to perform isomerization and reduction reactions, how these opposing pathways may co-exist within the same compartment and why so many oxidoreductases exist when PDI alone can perform all three of these functions.

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