July 26, 2022
Release Subtitle:
Researchers design and trace a modified cholera toxin to cell organelles
for insights on glycoprotein trafficking inside compartments of cells
Release Summary Text:
How are sugar moieties added to proteins inside cells? Although
extensively studied, details of the mechanism involved in protein
glycosylation at the level of organelles—subcellular compartments—remain
fuzzy, owing to difficulties in visualizing the transport of
glycoproteins inside these organelles. To unravel the mystery,
researchers from Japan have now designed and synthesized an artificial
protein derived from the cholera toxin that can be monitored inside the
cell as it travels to cell organelles.
Full text of release:
Proteins usually undergo modifications during or after their synthesis
in the endoplasmic reticulum (ER) and Golgi apparatus network inside
eukaryotic cells. One such modification is glycosylation, whereby
sugars, such as glycans, are added to newly synthesized proteins.
Glycans allow proteins to fold properly, in turn making them stable and
biologically active for various cell processes. However, the exact
mechanism of glycosylation in the ER and Golgi are still not known. One
way to study the process of glycosylation during protein synthesis is to
deliver synthetic proteins to specific cell organelles and observe
their subcellular dynamics. But this is often hindered by the lack of
specific delivery methods to organelles like the ER and Golgi.
To this end, Dr. Ayano Satoh from Okayama University and Dr. Yuta Maki,
Kazuki Kawata, Dr. Yanbo Liu, Kang-Ying Goo, Dr. Ryo Okamoto, and Prof.
Dr. Yasuhiro Kajihara from Osaka University, Japan investigated the
feasibility of modifying cholera toxin (CT) for targeted delivery to the
ER and Golgi. CT is a protein produced by the bacterium Vibrio cholerae
and is responsible for the hallmark symptoms of diarrhea—repeated
loose, watery stools. The toxin is made up of two subunits: CTA, which
causes diarrhea, and CTB, which helps the toxin enter cells. CT enters
the cell through the membrane into small cellular vehicles called
endosomes that deliver it to the Golgi bodies. From there, an
ER-specific amino acid sequence of CTA takes CT into the ER, where the
toxin springs into action to cause diarrhea. “CT
is a protein that naturally gets delivered specifically to the Golgi
and ER. This made it an attractive candidate for our investigation,”
says Dr. Satoh, explaining the reason behind selecting this protein for
their study, which was first published on May 23, 2022, in Chemistry – A European Journal.
The team synthesized an artificial, glycosylated form of the non-toxic
CTB and tracked its intracellular journey using the HiBiT
bioluminescence system engineered from the luciferase enzyme. In the
system that the team used, the larger fragment of luciferase was added
to particular receptors on the ER and Golgi. CTB was tagged with the
smaller fragment of luciferase. The system works by emitting light when
the two fragments bind to each other. Thus, the team tracked the
artificial CTB’s movement through the organelles in real time by
checking for the emittance of light. Talking about the highlights of
their study, Dr. Satoh says, “We
designed and chemically synthesized the glycosyl-CTB and demonstrated
its trafficking into the ER and Golgi of living cells. We also
established a method to quantitatively monitor the trafficking of CTB to
these organelles.”
The successful monitoring and delivery of the artificial CTB may pave
the way for a new phase of research in understanding protein
modification in compartments of living cells. The team emphasizes that
their method of preparing CTB allows for developing various mutant forms
of the protein as well as CTB bearing different glycans on its surface
to help investigate the functions of N-glycan in cells.
Not only the study of glycans but CTB-mediated delivery can also be a
promising tool for target-specific drug delivery in cells and
organelles. Dr. Satoh observes, “Our
system for targeting specific organelles may help treat diseases caused
by the absence of enzymes localized in specific organelles.”
What is her vision for the future? “Current
drug delivery techniques are limited because they only target the cell
surface. Our system may extend the limits of current technology and
enable the delivery of drug wherever it is needed,” says a hopeful Dr. Satoh.
We have our fingers crossed for her vision to come true and revolutionize the field of medicine!
Release URL:
https://www.eurekalert.org/news-releases/959718
Reference:
Design and Synthesis of Glycosylated Cholera Toxin B Subunit as a Tracer
of Glycoprotein Trafficking in Organelles of Living Cells
Journal: Chemistry – A European Journal
DOI:10.1002/chem.202201253
Contact Person:Ayano Satoh
Dr. Ayano Satoh is an Associate Professor of the Graduate School of
Interdisciplinary Science and Engineering in Health Systems and the
Faculty of Engineering at the Okayama University, Okayama, Japan. She
obtained her Bachelor’s and Master’s degrees in Chemistry from
Ochanomizu University, Tokyo, Japan. She then received her PhD at the
Graduate School of Humanities and Sciences, Ochanomizu University, where
she worked on affinity interactions among lipids, glycans, and proteins
with Dr. Isamu Matsumoto. Before joining Okayama University, she worked
with Dr. Graham Warren at Yale University, first as a Postdoctoral
Researcher, and then as a Research Scientist. At Yale University, she
worked on transport within the Golgi apparatus.
https://www.okayama-u.ac.jp/eng/research_highlights/index_id165.html
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