scholarly journals A toolbox of nanobodies developed and validated for diverse neuroscience research applications

2019 ◽  
Author(s):  
Jie-Xian Dong ◽  
Yongam Lee ◽  
Michael Kirmiz ◽  
Stephanie Palacio ◽  
Camelia Dumitras ◽  
...  
Keyword(s):  
Biomedical Research ◽  
Mammalian Cells ◽  
Structure And Function ◽  
Mammalian Brain ◽  
Tissue Penetration ◽  
Brain Neurons ◽  
Neuroscience Research ◽  
Form Part ◽  
Specific Proteins ◽  
And Function

SUMMARYNanobodies (nAbs) are small, minimal antibodies that have distinct attributes that make them uniquely suited for certain biomedical research, diagnostic and therapeutic applications. Prominent uses include as intracellular antibodies or intrabodies to bind and deliver cargo to specific proteins and/or subcellular sites within cells, and as nanoscale immunolabels for enhanced tissue penetration and improved spatial imaging resolution. Here, we report the generation and validation of nAbs against a set of proteins prominently expressed at specific subcellular sites in brain neurons. We describe a novel hierarchical validation pipeline to systematically evaluate nAbs isolated by phage display for effective and specific use as intrabodies and immunolabels in mammalian cells including brain neurons. These nAbs form part of a robust toolbox for targeting proteins with distinct and highly spatially-restricted subcellular localization in mammalian brain neurons, allowing for visualization and/or modulation of structure and function at those sites.

scholarly journals A toolbox of nanobodies developed and validated for use as intrabodies and nanoscale immunolabels in mammalian brain neurons

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Jie-Xian Dong ◽  
Yongam Lee ◽  
Michael Kirmiz ◽  
Stephanie Palacio ◽  
Camelia Dumitras ◽  
...  
Keyword(s):  
Subcellular Localization ◽  
Biomedical Research ◽  
Mammalian Cells ◽  
Structure And Function ◽  
Mammalian Brain ◽  
Tissue Penetration ◽  
Brain Neurons ◽  
Form Part ◽  
Specific Proteins ◽  
And Function

Nanobodies (nAbs) are small, minimal antibodies that have distinct attributes that make them uniquely suited for certain biomedical research, diagnostic and therapeutic applications. Prominent uses include as intracellular antibodies or intrabodies to bind and deliver cargo to specific proteins and/or subcellular sites within cells, and as nanoscale immunolabels for enhanced tissue penetration and improved spatial imaging resolution. Here, we report the generation and validation of nAbs against a set of proteins prominently expressed at specific subcellular sites in mammalian brain neurons. We describe a novel hierarchical validation pipeline to systematically evaluate nAbs isolated by phage display for effective and specific use as intrabodies and immunolabels in mammalian cells including brain neurons. These nAbs form part of a robust toolbox for targeting proteins with distinct and highly spatially-restricted subcellular localization in mammalian brain neurons, allowing for visualization and/or modulation of structure and function at those sites.

Oxidative protein folding in the mammalian endoplasmic reticulum

2004 ◽  
Vol 32 (5) ◽  
pp. 655-658 ◽  
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.

Expanding the genetic code of mammalian cells

2017 ◽  
Vol 45 (2) ◽  
pp. 555-562 ◽  
Author(s):  
James S. Italia ◽  
Yunan Zheng ◽  
Rachel E. Kelemen ◽  
Sarah B. Erickson ◽  
Partha S. Addy ◽  
...  
Keyword(s):  
Protein Structure ◽  
Amino Acid ◽  
Mammalian Cells ◽  
Living Cells ◽  
Structure And Function ◽  
Unnatural Amino Acid ◽  
Full Potential ◽  
Recent Developments ◽  
Small Footprint ◽  
And Function

In the last two decades, unnatural amino acid (UAA) mutagenesis has emerged as a powerful new method to probe and engineer protein structure and function. This technology enables precise incorporation of a rapidly expanding repertoire of UAAs into predefined sites of a target protein expressed in living cells. Owing to the small footprint of these genetically encoded UAAs and the large variety of enabling functionalities they offer, this technology has tremendous potential for deciphering the delicate and complex biology of the mammalian cells. Over the last few years, exciting progress has been made toward expanding the toolbox of genetically encoded UAAs in mammalian cells, improving the efficiency of their incorporation and developing innovative applications. Here, we provide our perspective on these recent developments and highlight the current challenges that must be overcome to realize the full potential of this technology.

scholarly journals Drosophila MICOS knockdown impairs mitochondrial structure and function and promotes mitophagy in muscle tissue

Biology Open ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. bio054262
Author(s):  
Li-jie Wang ◽  
Tian Hsu ◽  
Hsiang-ling Lin ◽  
Chi-yu Fu
Keyword(s):  
Muscle Tissue ◽  
Mammalian Cells ◽  
Structure And Function ◽  
Tissue Homeostasis ◽  
Mitochondrial Morphology ◽  
Mitochondrial Structure ◽  
And Function ◽  
Nucleoid Structure

ABSTRACTThe mitochondrial contact site and cristae organizing system (MICOS) is a multi-protein interaction hub that helps define mitochondrial ultrastructure. While the functional importance of MICOS is mostly characterized in yeast and mammalian cells in culture, the contributions of MICOS to tissue homeostasis in vivo remain further elucidation. In this study, we examined how knocking down expression of Drosophila MICOS genes affects mitochondrial function and muscle tissue homeostasis. We found that CG5903/MIC26-MIC27 colocalizes and functions with Mitofilin/MIC60 and QIL1/MIC13 as a Drosophila MICOS component; knocking down expression of any of these three genes predictably altered mitochondrial morphology, causing loss of cristae junctions, and disruption of cristae packing. Furthermore, the knockdown flies exhibited low mitochondrial membrane potential, fusion/fission imbalances, increased mitophagy, and limited cell death. Reductions in climbing ability indicated deficits in muscle function. Knocking down MICOS genes also caused reduced mtDNA content and fragmented mitochondrial nucleoid structure in Drosophila. Together, our data demonstrate an essential role of Drosophila MICOS in maintaining proper homeostasis of mitochondrial structure and function to promote the function of muscle tissue.

The Structure and Function of the Melian Dialogue

1968 ◽  
Vol 88 ◽  
pp. 73-77 ◽  
Author(s):  
W. Liebeschuetz
Keyword(s):  
Structure And Function ◽  
The Other ◽  
Independent State ◽  
Complete Destruction ◽  
Other Hand ◽  
Form Part ◽  
And Function ◽  
The City

Thucydides' account of the events at Melos in 416–15 B.C. falls into two parts, the famous dialogue in which representatives of Melos and of Athens discuss the submission of Melos, and a series of notes about the siege of the city culminating in the account of its destruction. But as I shall try to show, the two sections form part of a single whole. The discussion between the negotiators centres on two topics. In the first half of the dialogue the speakers discuss the expediency of forcing Melos into the Athenian Empire, in the second they discuss the likelihood of the Melians resisting successfully. But since the Melians are offered no alternative to becoming subjects except complete destruction, and since they are clearly not ready to choose the safe but dishonouring alternative, even though they have no chance of defending their city successfully, the inevitable destruction of Melos casts its shadow over the whole of the negotiations.Many of the arguments used in the discussion are equally relevant to the destruction and to the subjection of Melos. This is partly a result of the Athenian aim: to impress their island-subjects with their power. This they can achieve by forcing Melos to become a subject—but equally well by destroying it. On the level of expediency the Athenian argument would be equally applicable to either course, and the tactlessness of the Athenians suggests that they are not much concerned which of the two they will adopt. The Melians on the other hand appear to anticipate their own rejection of the ultimatum and to include the consequences of this inevitable rejection within the scope of their arguments. So a debate about the expediency of forcing an independent state to forgo its freedom is at the same time a debate about the expediency of destroying an independent city that refuses to become a subject.

Analysis of Adhesion Plaque Structure and Function in the Mechanism of Transformation by Rous Sarcoma Virus

1982 ◽  
Vol 40 ◽  
pp. 110-113
Author(s):  
L.R. Rohrschneider ◽  
M.J. Rosok ◽  
L.E. Gentry
Keyword(s):  
Rous Sarcoma Virus ◽  
Mammalian Cells ◽  
Neoplastic Transformation ◽  
Structure And Function ◽  
Excellent Opportunity ◽  
Cells In Culture ◽  
Rous Sarcoma ◽  
Sarcoma Virus ◽  
And Function ◽  
Adhesion Plaque

Rous sarcoma virus (RSV) was originally isolated from a fibrosarcoma of a chicken. This virus also will efficiently infect and transform all avian cells in culture as well as most mammalian cells. The mechanism of transformation by RSV is therefore universal and this system offers an excellent opportunity to investigate the mechanism of neoplastic transformation.

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