Subcellular Localization of Transiently Expressed Fluorescent Fusion Proteins

2013 ◽  
pp. 227-258 ◽  
Author(s):  
David A. Collings
Keyword(s):  
Subcellular Localization ◽  
Fusion Proteins

scholarly journals Differential Subcellular Localization Regulates Oncogenic Signaling by ROS1 Kinase Fusion Proteins

2018 ◽  
Vol 79 (3) ◽  
pp. 546-556 ◽  
Author(s):  
Dana S. Neel ◽  
David V. Allegakoen ◽  
Victor Olivas ◽  
Manasi K. Mayekar ◽  
Golzar Hemmati ◽  
...  
Keyword(s):  
Subcellular Localization ◽  
Fusion Proteins ◽  
Oncogenic Signaling

A simple and effective method for protein subcellular localization using Agrobacterium-mediated transformation of onion epidermal cells

Biologia ◽  
2007 ◽  
Vol 62 (5) ◽  
Author(s):  
Wei Sun ◽  
Ziyi Cao ◽  
Yan Li ◽  
Yanxiu Zhao ◽  
Hui Zhang
Keyword(s):  
Subcellular Localization ◽  
Particle Bombardment ◽  
Fusion Proteins ◽  
Fluorescent Proteins ◽  
Transformed Cells ◽  
Protein Subcellular Localization ◽  
Transformation Protocol ◽  
Agrobacterium Mediated Transformation ◽  
The Mean ◽  
Onion Epidermal Cells

AbstractA modified Agrobacterium-mediated transformation protocol has been successfully used for transient expression of the intrinsically fluorescent proteins and their fusion proteins in onion epidermis. The mean of the transformed cells rate per peel is about 10.5±0.9%, while that of the particle bombardment method is at the range 2.0±0.4%. To compare with the prevailing method of micro-projectile bombardment, the modified Agrobacterium-mediated transformation may provide with higher efficiency and even more simplified manipulability on a lower budget.

scholarly journals The C-terminus of cytochrome b5 confers endoplasmic reticulum specificity by preventing spontaneous insertion into membranes

2007 ◽  
Vol 401 (3) ◽  
pp. 701-709 ◽  
Author(s):  
Matthew P. A. Henderson ◽  
Yeen Ting Hwang ◽  
John M. Dyer ◽  
Robert T. Mullen ◽  
David W. Andrews
Keyword(s):  
Endoplasmic Reticulum ◽  
Subcellular Localization ◽  
Lipid Bilayers ◽  
Fusion Proteins ◽  
Molecular Mechanisms ◽  
Cytochrome B5 ◽  
Terminal Sequence ◽  
C Terminus

The molecular mechanisms that determine the correct subcellular localization of proteins targeted to membranes by tail-anchor sequences are poorly defined. Previously, we showed that two isoforms of the tung oil tree [Vernicia (Aleurites) fordii] tail-anchored Cb5 (cytochrome b5) target specifically to ER (endoplasmic reticulum) membranes both in vivo and in vitro [Hwang, Pelitire, Henderson, Andrews, Dyer and Mullen (2004) Plant Cell 16, 3002–3019]. In the present study, we examine the targeting of various tung Cb5 fusion proteins and truncation mutants to purified intracellular membranes in vitro in order to assess the importance of the charged CTS (C-terminal sequence) in targeting to specific membranes. Removal of the CTS from tung Cb5 proteins resulted in efficient binding to both ER and mitochondria. Results from organelle competition, liposome-binding and membrane proteolysis experiments demonstrated that removal of the CTS results in spontaneous insertion of tung Cb5 proteins into lipid bilayers. Our results indicate that the CTSs from plant Cb5 proteins provide ER specificity by preventing spontaneous insertion into incorrect subcellular membranes.

Subcellular Localization of Signal Peptide Fusion Proteins Expressed in E. coli

2021 ◽  
Vol 2021 (2) ◽  
pp. pdb.prot102145
Author(s):  
Clara L. Kielkopf ◽  
William Bauer ◽  
Ina L. Urbatsch
Keyword(s):  
Subcellular Localization ◽  
Signal Peptide ◽  
Fusion Proteins ◽  
E Coli

Analysis of Phosphoinositide-Binding Properties and Subcellular Localization of GFP-Fusion Proteins

Lipids ◽  
2015 ◽  
Vol 50 (4) ◽  
pp. 427-436 ◽  
Author(s):  
Yong-Woo Jun ◽  
Sangyeol Kim ◽  
Kun-Hyung Kim ◽  
Jin-A Lee ◽  
Chae-Seok Lim ◽  
...  
Keyword(s):  
Subcellular Localization ◽  
Fusion Proteins ◽  
Binding Properties

802 Subcellular localization of GFP-drebrin fusion proteins

1997 ◽  
Vol 28 ◽  
pp. S100
Author(s):  
Kensuke Hayashi ◽  
Tomoaki Shirao
Keyword(s):  
Subcellular Localization ◽  
Fusion Proteins

scholarly journals Investigation Into the use of C- and N-terminal GFP Fusion Proteins for Subcellular Localization Studies Using Reverse Transfection Microarrays

2004 ◽  
Vol 5 (4) ◽  
pp. 342-353 ◽  
Author(s):  
Ella Palmer ◽  
Tom Freeman
Keyword(s):  
Subcellular Localization ◽  
High Throughput ◽  
Fusion Proteins ◽  
Fluorescent Protein ◽  
Protein Localization ◽  
Genes Encoding ◽  
Reverse Transfection ◽  
C And N ◽  
Green Fluorescent ◽  
Cellular Compartments

Reverse transfection microarrays were described recently as a high throughput method for studying gene function. We have investigated the use of this technology for determining the subcellular localization of proteins. Genes encoding 16 proteins with a variety of functions were placed in Gateway expression constructs with 3′ or 5′ green fluorescent protein (GFP) tags. These were then packaged in transfection reagent and spotted robotically onto a glass slide to form a reverse transfection array. HEK293T cells were grown over the surface of the array until confluent and GFP fluorescence visualized by confocal microscopy. All C-terminal fusion proteins localized to cellular compartments in accordance with previous studies and/or bioinformatic predictions. However, less than half of the N-terminal fusion proteins localized correctly. Of those that were not in concordance with the C-terminal tagged proteins, half did not exhibit expression and the remainder had differing subcellular localizations to the C-terminal fusion protein. This data indicates that N-terminal tagging with GFP adversely affects the protein localization in reverse transfection assays, whereas tagging with GFP at the C-terminal is generally better in preserving the localization of the native protein. We discuss these results in the context of developing high-throughput subcellular localization assays based on the reverse transfection array technology.

Sign in / Sign up

Export Citation Format

Share Document