scholarly journals Cellular Localization of Exogenous Cry1Ab/c and its Interaction with Plasma Membrane Ca2+-ATPase in Transgenic Rice

2021 ◽  
Vol 9 ◽  
Author(s):  
Jianmei Fu ◽  
Yu Shi ◽  
Laipan Liu ◽  
Biao Liu
Keyword(s):  
Plasma Membrane ◽  
Salt Stress ◽  
Membrane Protein ◽  
Atpase Activity ◽  
Transgenic Rice ◽  
Growth And Development ◽  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Plasma Membrane Protein ◽  
C Protein

The cellular localization of exogenous proteins expressed in transgenic crops not only determines their stability, but also their effects on crop growth and development, including under stressful conditions; however, the underlying molecular mechanisms remain unknown. Here, we determined the cellular distribution of exogenously expressed Cry1Ab/c protein in insect-resistant transgenic rice Huahui-1 (HH1) cells through subcellular localization, immunohistochemistry, immunofluorescence, and western blot analyses. Interaction between the Cry1Ab/c protein and the preliminarily screened endogenous plasma membrane protein Ca2+-ATPase was investigated through yeast two-hybrid, bimolecular fluorescence complementation (BIFC), and co-immunoprecipitation analyses. The potential interaction mechanism was analyzed by comparing the cellular localization and interaction sites between Cry1Ab/c and Ca2+-ATPase. Phenotypic indices and Ca2+-ATPase activity, which may be regulated by the Cry1Ab/c–Ca2+-ATPase interaction, were determined in transgenic HH1 and the parental line Minghui-63 under stress-free and salt-stress conditions. The results showed that Cry1Ab/c was not only distributed in the cytoplasm and nucleus but was also distributed on the plasma membrane, where it interacted with plasma membrane Ca2+-ATPase. This interaction partially retain plasma membrane protein Ca2+-ATPase in the nucleus by a BIFC experiment and thus may affect Ca2+-ATPase activity on the membrane by altering the cellular location of the protein. Consistently, our results confirmed that the presence of Cry1Ab/c in the transgenic HH1 resulted in a reduction in Ca2+-ATPase activity as well as causing detrimental effects on plant phenotype, including significantly reduced plant height and biomass, compared to parental MH63; and that these detrimental effects were more pronounced under salt stress conditions, impacting the salt resistance of the transgenic plants. We suggest that the Cry1Ab/c–Ca2+-ATPase interaction may explain the plasma membrane localization of Cry1Ab/c, which lacks a signal peptide and a transmembrane domain, and the adverse effects of Cry1Ab/c expression on the growth and development of transgenic HH1 plants under salt stress. This information may clarify the molecular mechanisms of these unintended effects and demonstrate the feasibility of evaluating the success and performance of genetic modification of commercially vital crops.

Functional expression and cellular localization of the Na+/H+ exchanger Sod2 of the fission yeast Schizosaccharomyces pombe

2005 ◽  
Vol 83 (7) ◽  
pp. 565-572 ◽  
Author(s):  
Larry Fliegel ◽  
Christine Wiebe ◽  
Gordon Chua ◽  
Paul G Young
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Cellular Localization ◽  
Functional Expression ◽  
Cation Binding ◽  
Plasma Membrane Protein ◽  
Gfp Fusion Protein ◽  
Growing Cell

In the fission yeast Schizosaccharomyces pombe, the Na+/H+ exchanger, Sod2, plays a major role in the removal of excess intracellular sodium, and its disruption results in a sodium-sensitive phenotype. We examined the subcellular distribution and dynamics of Sod2 expression in S. pombe using a sod2-GFP fusion protein under the control of an attenuated version of the inducible nmt promoter. Sod2 was localized throughout the plasma membrane, the nuclear envelope, and some internal membrane systems. In exponentially growing cells, in which sod2-GFP was expressed and then the promoter turned-off, previously synthesized sod2-GFP was stable for long periods and found localized to the plasma membrane in the medial regions of the cell. It was not present at the actively growing cell ends. This suggests that these regions of the cell contain old plasma membrane protein vs. newly synthesized plasma membrane without Sod2 at the growing ends. Sod2 localization was not affected by salt stress. The results suggest that Sod2 is both a plasma membrane protein and is present in intracellular membranes. It is likely tethered within discrete regions of the plasma membrane and is not free to diffuse throughout the bilayer. Key words: Na+/H+ exchanger, Schizosaccharomyces pombe, cation binding, salt tolerance.

scholarly journals The 3'-UTR mediates the cellular localization of an mRNA encoding a short plasma membrane protein

RNA ◽  
2008 ◽  
Vol 14 (7) ◽  
pp. 1352-1365 ◽  
Author(s):  
A. Loya ◽  
L. Pnueli ◽  
Y. Yosefzon ◽  
Y. Wexler ◽  
M. Ziv-Ukelson ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Cellular Localization ◽  
Plasma Membrane Protein

The transcytotic pathway of an apical plasma membrane protein (B10) in hepatocytes is similar to that of IgA and occurs via a tubular pericentriolar compartment

1996 ◽  
Vol 109 (6) ◽  
pp. 1215-1227 ◽  
Author(s):  
I. Hemery ◽  
A.M. Durand-Schneider ◽  
G. Feldmann ◽  
J.P. Vaerman ◽  
M. Maurice
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Apical Membrane ◽  
Rat Hepatocytes ◽  
Apical Plasma Membrane ◽  
Apical Surface ◽  
Plasma Membrane Protein ◽  
Microtubule Disruption

In hepatocytes, newly synthesized apical plasma membrane proteins are first delivered to the basolateral surface and are supposed to reach the apical surface by transcytosis. The transcytotic pathway of apical membrane proteins and its relationship with other endosomal pathways has not been demonstrated morphologically. We compared the intracellular route of an apical plasma membrane protein, B10, with that of polymeric IgA (pIgA), which is transcytosed, transferrin (Tf) which is recycled, and asialoorosomucoid (ASOR) which is delivered to lysosomes. Ligands and anti-B10 monoclonal IgG were linked to fluorochromes or with peroxidase. The fate of each ligand was followed by confocal and electron microscopy in polarized primary monolayers of rat hepatocytes. When fluorescent anti-B10 IgG and fluorescent pIgA were simultaneously endocytosed for 15–30 minutes, they both uniformly labelled a juxtanuclear compartment. By 30–60 minutes, they reached the bile canaliculi. Tf and ASOR were also routed to the juxtanuclear area, but their fluorescence patterns were more punctate. Microtubule disruption prevented all ligands from reaching the juxtanuclear area. This area corresponded, at least partially, to the localization of the mannose 6-phosphate receptor, an endosomal marker. By electron microscopy, the juxtanuclear compartment was made up of anastomosing tubules connected to vacuoles, and was organized around the centrioles. B10 and pIgA were mainly found in the tubules, whereas ASOR was segregated inside the vacuolar elements and Tf within thinner, recycling tubules. In conclusion, transcytosis of the apical membrane protein B10 occurs inside tubules similar to those carrying pIgA, and involves passage via the pericentriolar area. In the pericentriolar area, the transcytotic tubules appear to maintain connections with other endosomal elements where sorting between recycled and degraded ligands occurs.

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