The Experts below are selected from a list of 303 Experts worldwide ranked by ideXlab platform
Tom A Rapoport - One of the best experts on this subject based on the ideXlab platform.
-
the Signal Sequence coding region promotes nuclear export of mrna
PLOS Biology, 2007Co-Authors: Alexander F Palazzo, Michael Springer, Yoko Shibata, Chungsheng Lee, Anusha P Dias, Tom A RapoportAbstract:In eukaryotic cells, most mRNAs are exported from the nucleus by the transcription export (TREX) complex, which is loaded onto mRNAs after their splicing and capping. We have studied in mammalian cells the nuclear export of mRNAs that code for secretory proteins, which are targeted to the endoplasmic reticulum membrane by hydrophobic Signal Sequences. The mRNAs were injected into the nucleus or synthesized from injected or transfected DNA, and their export was followed by fluorescent in situ hybridization. We made the surprising observation that the Signal Sequence coding region (SSCR) can serve as a nuclear export Signal of an mRNA that lacks an intron or functional cap. Even the export of an intron-containing natural mRNA was enhanced by its SSCR. Like conventional export, the SSCR-dependent pathway required the factor TAP, but depletion of the TREX components had only moderate effects. The SSCR export Signal appears to be characterized in vertebrates by a low content of adenines, as demonstrated by genome-wide Sequence analysis and by the inhibitory effect of silent adenine mutations in SSCRs. The discovery of an SSCR-mediated pathway explains the previously noted amino acid bias in Signal Sequences and suggests a link between nuclear export and membrane targeting of mRNAs.
-
Signal Sequence recognition in posttranslational protein transport across the yeast er membrane
Cell, 1998Co-Authors: Kathrin Plath, Walther Mothes, Barrie M Wilkinson, Colin J Stirling, Tom A RapoportAbstract:Abstract We have analyzed how the Signal Sequence of prepro- α -factor is recognized during the first step of posttranslational protein transport into the yeast endoplasmic reticulum. Cross-linking studies indicate that the Signal Sequence interacts in a Kar2p- and ATP-independent reaction with Sec61p, the multispanning membrane component of the protein-conducting channel, by intercalation into transmembrane domains 2 and 7. While bound to Sec61p, the Signal Sequence forms a helix that is contacted on one side by Sec62p and Sec71p. The binding site is located at the interface of the protein channel and the lipid bilayer. Signal Sequence recognition in cotranslational translocation in mammals appears to occur similarly. These results suggest a general mechanism by which the Signal Sequence could open the channel for polypeptide transport.
-
Signal Sequence dependent function of the tram protein during early phases of protein transport across the endoplasmic reticulum membrane
Journal of Cell Biology, 1996Co-Authors: Sabine Voigt, Berit Jungnickel, E Hartmann, Tom A RapoportAbstract:Cotranslational translocation of proteins across the mammalian ER membrane involves, in addition to the Signal recognition particle receptor and the Sec61p complex, the translocating chain-associating membrane (TRAM) protein, the function of which is still poorly understood. Using reconstituted proteoliposomes, we show here that the translocation of most, but not all, secretory proteins requires the function of TRAM. Experiments with hybrid proteins demonstrate that the structure of the Signal Sequence determines whether or not TRAM is needed. Features that distinguish TRAM-dependent and -independent Signal Sequences include the length of their charged, NH2-terminal region and the structure of their hydrophobic core. In cases where TRAM is required for translocation, it is not needed for the initial interaction of the ribosome/nascent chain complex with the ER membrane but for a subsequent step inside the membrane in which the nascent chain is inserted into the translocation site in a protease-resistant manner. Thus, TRAM functions in a Signal Sequence-dependent manner at a critical, early phase of the translocation process.
-
a posttargeting Signal Sequence recognition event in the endoplasmic reticulum membrane
Cell, 1995Co-Authors: Berit Jungnickel, Tom A RapoportAbstract:Abstract We have analyzed early phases of the cotranslational transport of the secretory protein preprolactin through the mammalian endoplasmic reticulum (ER) membrane. Following recognition of the Signal Sequence of the nascent polypeptide chain in the cytosol by the SRP, the chain is transferred into the membrane, where a second Signal Sequence recognition step takes place for which the presence in the lipid bilayer of the Sec61 p complex is essential and sufficient. This step leads to a tight junction between the ribosomenascent chain complex and the Sec61p complex, and to the productive insertion of the nascent chain into the translocation site. These results show that a translocation substrate is subjected to two recognition events before being allowed to cross the ER membrane.
-
site specific photocross linking reveals that sec61p and tram contact different regions of a membrane inserted Signal Sequence
Journal of Biological Chemistry, 1993Co-Authors: Stephen High, Tom A Rapoport, Bruno Martoglio, Dirk Gorlich, S S Andersen, Anthony J Ashford, Angelika Giner, Enno Hartmann, Siegfried Prehn, Bernhard DobbersteinAbstract:A chemically charged amber suppressor tRNA was used to introduce the photoactivatable amino acid (Tmd)Phe at a selected position within the Signal Sequence of the secretory protein preprolactin. This allowed the interactions of the NH2-terminal, the central, and the COOH-terminal regions of the Signal Sequence to be investigated during insertion into the membrane of the endoplasmic reticulum (ER). We found that different regions of the nascent chains were photocross-linked to different ER proteins. The TRAM protein (translocating chain-associating membrane protein) contacts the NH2-terminal region of the Signal Sequence while the mammalian Sec61p contacts the hydrophobic core of the Signal Sequence and regions COOH-terminal of this. These results suggest that the ER translocation complex is composed of heterologous protein subunits which contact distinct regions of nascent polypeptides during their membrane insertion.
Evalina Williamson - One of the best experts on this subject based on the ideXlab platform.
-
a putative nuclear nucleolar Signal Sequence directs the localization of trpm7 kinase
Biophysical Journal, 2017Co-Authors: Ceredwyn E Hill, Adenike Ogunrinde, Christiane Whetstone, Evalina WilliamsonAbstract:The channel-kinase TRPM7 supports the survival, proliferation, and differentiation of many cell types. Both plasma membrane channel activity and kinase function have been implicated in these roles, although the specific functions of either have not been clearly established. The kinase has been invoked in the regulation of protein synthesis, chromatin status, actomyosin contractility, allergy and inflammation. Identified proteinaceous substrates are localized throughout the cytoplasm and nucleus, lending credence to the observation that the soluble kinase domain can be cleaved from the channel (Krapivinsky et al. 2014). The same study reported that a kinase construct (1510-1863), heterologously expressed in HEK293 cells, appears in the nucleus. We recently showed that hepatoma cells express a soluble C-terminal immunoreactive domain of TRPM7 in the nucleus and nucleolus, whereas non-dividing hepatocytes express nuclear envelope but not nucleoplasmic, chanzyme. To further our understanding of the cellular role(s) of the kinase domain and establish how the kinase is targeted to specific nuclear compartments, we used genetic and immunofluorescence approaches to identify the subcellular location(s) of TRPM7 kinase constructs containing putative subcellular localization Sequences. HEK293T cells heterologously expressing the isolated kinase domain (1510-1863) confirmed the nucleoplasmic localization. Contrary to the endogenous labelling in hepatoma, this protein did not appear in the nucleolus. However, a larger construct containing a putative nucleolar Signalling Sequence (K1147-K1151) labelled the nucleoplasm and the nucleolus, where it colocalizes with fibrillarin. Western blotting of fractionated nuclei confirmed these results. Thus the TRPM7 kinase appears to take different forms, possibly cell specific, allowing it to localize to specific nuclear locations. We surmise that TRPM7 and its cleavage products distribute between the plasma membrane and nucleus in proliferating hepatoma cells. Nucleolar kinase activity may be involved in nucleolar functions such as ribosome synthesis.
-
a putative nuclear nucleolar Signal Sequence directs the localization of trpm7 kinase
Biophysical Journal, 2017Co-Authors: Ceredwyn E Hill, Adenike Ogunrinde, Christiane Whetstone, Evalina WilliamsonAbstract:The channel-kinase TRPM7 supports the survival, proliferation, and differentiation of many cell types. Both plasma membrane channel activity and kinase function have been implicated in these roles, although the specific functions of either have not been clearly established. The kinase has been invoked in the regulation of protein synthesis, chromatin status, actomyosin contractility, allergy and inflammation. Identified proteinaceous substrates are localized throughout the cytoplasm and nucleus, lending credence to the observation that the soluble kinase domain can be cleaved from the channel (Krapivinsky et al. 2014). The same study reported that a kinase construct (1510-1863), heterologously expressed in HEK293 cells, appears in the nucleus. We recently showed that hepatoma cells express a soluble C-terminal immunoreactive domain of TRPM7 in the nucleus and nucleolus, whereas non-dividing hepatocytes express nuclear envelope but not nucleoplasmic, chanzyme. To further our understanding of the cellular role(s) of the kinase domain and establish how the kinase is targeted to specific nuclear compartments, we used genetic and immunofluorescence approaches to identify the subcellular location(s) of TRPM7 kinase constructs containing putative subcellular localization Sequences. HEK293T cells heterologously expressing the isolated kinase domain (1510-1863) confirmed the nucleoplasmic localization. Contrary to the endogenous labelling in hepatoma, this protein did not appear in the nucleolus. However, a larger construct containing a putative nucleolar Signalling Sequence (K1147-K1151) labelled the nucleoplasm and the nucleolus, where it colocalizes with fibrillarin. Western blotting of fractionated nuclei confirmed these results. Thus the TRPM7 kinase appears to take different forms, possibly cell specific, allowing it to localize to specific nuclear locations. We surmise that TRPM7 and its cleavage products distribute between the plasma membrane and nucleus in proliferating hepatoma cells. Nucleolar kinase activity may be involved in nucleolar functions such as ribosome synthesis.
Ceredwyn E Hill - One of the best experts on this subject based on the ideXlab platform.
-
a putative nuclear nucleolar Signal Sequence directs the localization of trpm7 kinase
Biophysical Journal, 2017Co-Authors: Ceredwyn E Hill, Adenike Ogunrinde, Christiane Whetstone, Evalina WilliamsonAbstract:The channel-kinase TRPM7 supports the survival, proliferation, and differentiation of many cell types. Both plasma membrane channel activity and kinase function have been implicated in these roles, although the specific functions of either have not been clearly established. The kinase has been invoked in the regulation of protein synthesis, chromatin status, actomyosin contractility, allergy and inflammation. Identified proteinaceous substrates are localized throughout the cytoplasm and nucleus, lending credence to the observation that the soluble kinase domain can be cleaved from the channel (Krapivinsky et al. 2014). The same study reported that a kinase construct (1510-1863), heterologously expressed in HEK293 cells, appears in the nucleus. We recently showed that hepatoma cells express a soluble C-terminal immunoreactive domain of TRPM7 in the nucleus and nucleolus, whereas non-dividing hepatocytes express nuclear envelope but not nucleoplasmic, chanzyme. To further our understanding of the cellular role(s) of the kinase domain and establish how the kinase is targeted to specific nuclear compartments, we used genetic and immunofluorescence approaches to identify the subcellular location(s) of TRPM7 kinase constructs containing putative subcellular localization Sequences. HEK293T cells heterologously expressing the isolated kinase domain (1510-1863) confirmed the nucleoplasmic localization. Contrary to the endogenous labelling in hepatoma, this protein did not appear in the nucleolus. However, a larger construct containing a putative nucleolar Signalling Sequence (K1147-K1151) labelled the nucleoplasm and the nucleolus, where it colocalizes with fibrillarin. Western blotting of fractionated nuclei confirmed these results. Thus the TRPM7 kinase appears to take different forms, possibly cell specific, allowing it to localize to specific nuclear locations. We surmise that TRPM7 and its cleavage products distribute between the plasma membrane and nucleus in proliferating hepatoma cells. Nucleolar kinase activity may be involved in nucleolar functions such as ribosome synthesis.
-
a putative nuclear nucleolar Signal Sequence directs the localization of trpm7 kinase
Biophysical Journal, 2017Co-Authors: Ceredwyn E Hill, Adenike Ogunrinde, Christiane Whetstone, Evalina WilliamsonAbstract:The channel-kinase TRPM7 supports the survival, proliferation, and differentiation of many cell types. Both plasma membrane channel activity and kinase function have been implicated in these roles, although the specific functions of either have not been clearly established. The kinase has been invoked in the regulation of protein synthesis, chromatin status, actomyosin contractility, allergy and inflammation. Identified proteinaceous substrates are localized throughout the cytoplasm and nucleus, lending credence to the observation that the soluble kinase domain can be cleaved from the channel (Krapivinsky et al. 2014). The same study reported that a kinase construct (1510-1863), heterologously expressed in HEK293 cells, appears in the nucleus. We recently showed that hepatoma cells express a soluble C-terminal immunoreactive domain of TRPM7 in the nucleus and nucleolus, whereas non-dividing hepatocytes express nuclear envelope but not nucleoplasmic, chanzyme. To further our understanding of the cellular role(s) of the kinase domain and establish how the kinase is targeted to specific nuclear compartments, we used genetic and immunofluorescence approaches to identify the subcellular location(s) of TRPM7 kinase constructs containing putative subcellular localization Sequences. HEK293T cells heterologously expressing the isolated kinase domain (1510-1863) confirmed the nucleoplasmic localization. Contrary to the endogenous labelling in hepatoma, this protein did not appear in the nucleolus. However, a larger construct containing a putative nucleolar Signalling Sequence (K1147-K1151) labelled the nucleoplasm and the nucleolus, where it colocalizes with fibrillarin. Western blotting of fractionated nuclei confirmed these results. Thus the TRPM7 kinase appears to take different forms, possibly cell specific, allowing it to localize to specific nuclear locations. We surmise that TRPM7 and its cleavage products distribute between the plasma membrane and nucleus in proliferating hepatoma cells. Nucleolar kinase activity may be involved in nucleolar functions such as ribosome synthesis.
Mark R Payne - One of the best experts on this subject based on the ideXlab platform.
-
a novel tat mitochondrial Signal Sequence fusion protein is processed stays in mitochondria and crosses the placenta
Molecular Therapy, 2003Co-Authors: Victoria Del Gaizo, Mark R PayneAbstract:Mutations in nuclear and mitochondrial genomes can lead to defects in mitochondrial function. To date, repair of these defects with exogenous proteins or gene transfer has been difficult with either viral or nonviral vectors. We hypothesized that TAT fusion proteins would cross both mitochondrial membranes and that incorporation of a mitochondrial Signal Sequence into a TAT fusion protein would allow processing and localization of exogenous proteins in mitochondria. A TAT–mitochondrial malate dehydrogenase Signal Sequence (mMDH)–enhanced green fluorescent protein (eGFP) fusion protein was constructed. TAT-mMDH-eGFP allowed rapid transduction and localization of fusion protein into mitochondria of multiple cell types. In contrast, TAT-GFP, without a mitochondrial Signal Sequence, rapidly transduced into cells and mitochondria, displayed pseudo-first-order kinetics, but did not remain there. Mice injected 5 days prior with TAT-mMDH-eGFP had detectable eGFP activity in multiple tissue types. Western blotting of cytosolic and mitochondrial fractions isolated from their livers confirmed eGFP localization to mitochondria and that the mMDH transit peptide was recognized and processed. Furthermore, TAT-mMDH-eGFP fusion protein injected into pregnant mice crossed the placenta and was detectable in both the fetus and the newborn pups. TAT fusion proteins containing a mitochondrial Signal Sequence are a viable method to localize proteins to mitochondria.
Francois Sigaux - One of the best experts on this subject based on the ideXlab platform.
-
cutting edge requirement of class i Signal Sequence derived peptides for hla e recognition by a mouse cytotoxic t cell clone
Journal of Immunology, 1999Co-Authors: Silvia Martinozzi, Matthias Ulbrecht, Elisabeth H Weiss, Rita Pacasova, Henrijean Boulouis, Francois SigauxAbstract:The human nonclassical MHC class I molecule HLA-E has recently been shown to act as a major ligand for NK cell inhibitory receptors. Using HLA-E-expressing transgenic mice, we produced a cytotoxic T cell clone that specifically recognizes the HLA-E molecule. We report here that this T cell clone lyses HLA-E-transfected RMA-S target cells sensitized with synthetic class I Signal Sequence nonamers. Moreover, this T cell clone lyses human EBV-infected B lymphocytes, PHA blasts, and PBL, formally demonstrating the surface expression of HLA-E/class I Signal-derived peptide complex on human cells. Furthermore, these data show that HLA-E complexed with class I Signal Sequence-derived peptides is not only a ligand for NK cell inhibitory receptors, but can also trigger cytotoxic T cells (CTL).
