The complex process of GETting tail-anchored membrane proteins to the ER

Chartron, J.W.; Clemons, William M.; Suloway, Christian

doi:10.1016/j.sbi.2012.03.001

Cited by 69 publications

(103 citation statements)

References 42 publications

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“…In mammals, Bag6 has been shown to be critical in the targeting of TA proteins to the ER by the transmembrane recognition complex (TRC) pathway (5), a process best understood in the equivalent fungal guided entry of tail-anchored proteins (GET) pathway (17,18). Although Bag6 is missing in fungi, the analogous yeast complex contains two proteins, Get4 and Get5/ Mdy2, which are homologs of the mammalian proteins TRC35 and Ubl4A, respectively.…”

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confidence: 99%

Bag6 complex contains a minimal tail-anchor–targeting module and a mock BAG domain

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Chartron

Zaslaver

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

129

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BCL2-associated athanogene cochaperone 6 (Bag6) plays a central role in cellular homeostasis in a diverse array of processes and is part of the heterotrimeric Bag6 complex, which also includes ubiquitin-like 4A (Ubl4A) and transmembrane domain recognition complex 35 (TRC35). This complex recently has been shown to be important in the TRC pathway, the mislocalized protein degradation pathway, and the endoplasmic reticulum-associated degradation pathway. Here we define the architecture of the Bag6 complex, demonstrating that both TRC35 and Ubl4A have distinct C-terminal binding sites on Bag6 defining a minimal Bag6 complex. A crystal structure of the Bag6–Ubl4A dimer demonstrates that Bag6–BAG is not a canonical BAG domain, and this finding is substantiated biochemically. Remarkably, the minimal Bag6 complex defined here facilitates tail-anchored substrate transfer from small glutamine-rich tetratricopeptide repeat-containing protein α to TRC40. These findings provide structural insight into the complex network of proteins coordinated by Bag6.

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confidence: 99%

Bag6 complex contains a minimal tail-anchor–targeting module and a mock BAG domain

Mock

Chartron

Zaslaver

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

129

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“…6, with the emerging view that the overall hydropathy of the TMD plays an important role in determining pathway specificity 48 . From high to low overall hydropathy, the tail anchor can be targeted to the ER by the classical SRP pathway 13 , the Sec62/63 pathway 14 , the GET/TRC40 pathway [15][16][17][18][19][20][21][22] , more general chaperones [23][24][25] , or spontaneous insertion 27 . For all of these pathways, the Sec61 translocon likely plays either an essential or important role (even for spontaneous insertion).…”

Section: Investigation Of Tail Anchor Targeting Pathways That May Regmentioning

confidence: 99%

“…A major mechanism for tail insertion is through the GET/TRC40 pathway [15][16][17][18][19]21,22 . To examine its role in the insertion of the tail anchor of PTP1B, we systematically deleted proteins involved in this pathway in a yeast strain chromosomally expressing yemCitrine-PTP1Btail.…”

Section: Investigation Of Tail Anchor Targeting Pathways That May Regmentioning

confidence: 99%

“…PTP1B's short (≤35 amino acid) C-terminal tail anchor was previously reported to localize it on the endoplasmic reticulum (ER) 10,12 . Exactly how the tail anchor is inserted into the membrane of the ER remains unknown, with possible contributions from the classical signal recognition particle (SRP) pathway 13 , the Sec62/63 pathway 14 , the guided entry of tail anchor proteins (GET/TRC40) pathway [15][16][17][18][19][20][21][22] , chaperone-assisted interaction [23][24][25] with the ER translocon Sec61 26 , or through spontaneous insertion 27 . How these different pathways might act on PTP1B as well as how these pathways are generally tailored to engage the wide spectrum of tail-anchor-containing proteins remains largely unknown.…”

Section: Introductionmentioning

confidence: 99%

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Journey to the Center of the Mitochondria Guided by the Tail Anchor of Protein Tyrosine Phosphatase 1B

Fueller

Egorov

et al. 2013

Preprint

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The canonical protein tyrosine phosphatase PTP1B has traditionally been considered to exclusively reside on the endoplasmic reticulum (ER). Using confocal microscopy, we show that endogenous PTP1B actually exhibits a higher local concentration at the mitochondria in all mammalian cell lines that we tested. Fluorescently labeled chimeras containing full-length PTP1B or only its 35 amino acid tail anchor localized identically, demonstrating the complete dependence of PTP1B's subcellular partitioning on its tail anchor. Correlative light and electron microscopy using GFPdriven photo-oxidation of DAB revealed that PTP1B's tail anchor localizes it to the mitochondrial interior and to mitochondrial-associated membrane (MAM) sites along the ER. Heterologous expression of the tail anchor of PTP1B in the yeast S. cerevisiae surprisingly led to its exclusive localization to the ER/vacuole with no presence at the mitochondria. Studies with various yeast mutants of conserved membrane insertion pathways revealed a role for the GET/TRC40 pathway in ER insertion, but also emphasized the likely dominant role of spontaneous insertion. Further studies of modified tail isoforms in both yeast and mammalian cells revealed a remarkable sensitivity of subcellular partitioning to slight changes in transmembrane domain (TMD) length, C-terminal charge, and hydropathy. For example, addition of a single positive charge to the tail anchor was sufficient to completely shift the tail anchor to the mitochondria in mammalian cells and to largely shift it there in yeast cells, and a point mutation that increased TMD hydropathy was sufficient to localize the tail anchor exclusively to the ER in mammalian cells. Striking differences in the subcellular partitioning of a given tail anchor isoform in mammalian versus yeast cells most likely point to fundamental differences in the lipid composition of specific organelles (e.g. affecting membrane charge or thickness) in higher versus lower eukaryotes. Fluorescence lifetime imaging microscopy (FLIM) detection of the Förster Resonance Energy Transfer (FRET)-based interaction of the catalytic domain of PTP1B with the epidermal growth factor receptor (EGFR/ErbB1) at the mitochondria revealed a strong interaction on the cytosolic face of the outer mitochondrial membrane (OMM), suggesting the presence of a significant pool of PTP1B there and a novel role for PTP1B in the regulation of mitochondrial ErbB1 activity. In summary, in addition to its well-established general localization along the ER, our results reveal that PTP1B specifically accumulates at MAM sites along the ER and localizes as well to the OMM and mitochondrial matrix. Further elucidation of PTP1B's roles in these different locations (including the identification of its targets) will likely be critical for understanding its complex regulation of general cellular responses, cell proliferation, and diseased states.

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“…TAs, defined by a single transmembrane domain (TMD) near the C terminus, comprise up to 5% of the eukaryotic membrane proteome and mediate diverse key cellular functions, including protein translocation across multiple organelle membranes, vesicular fusion, protein quality control, and apoptosis (3). In eukaryotic cells, TAs are targeted to the endoplasmic reticulum (ER) by the conserved guided entry of tail-anchored protein (GET) pathway, in which the Get3 ATPase captures TAs with help of the cytosolic Get4/5 complex and then delivers TAs to the Get1/2 receptor complex at the ER membrane (4)(5)(6). During the targeting cycle, Get3 undergoes extensive changes in conformation and activity in response to nucleotides, effector proteins, and the TA substrate (Fig.…”

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confidence: 99%