Technion – Israel Institute of Technology
Department of Biotechnology & Food Engineering
Project name: Discover the location of iron storage
protein (ferritin) in macrophage cells
Scientific name: Elucidating ferritin localization in
murine bone marrow derived macrophages
Instructor: Marianna Truman (Ph.D. research
Department of Biotechnology and Food Engineering
Supervisor: Prof. Esther Meyron-Holtz,
Biotechnology and Food Engineering
student),
Department
of
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Abstract: Ferritin is a well-known iron storage and detoxification
protein. Apo-ferritin is a nearly spherical protein shell, and in
vertebrates it is composed of 24 subunits
mostly of two kinds, H- and L-ferritin,
arranged around a large cavity. Ferritin
has the ability to store up to 4,500 iron
atoms in a soluble, non-toxic and
biologically non-reactive form, and to
release iron when there is an increase in
Figure 1: Ferritin protein structure
the cell’s need for bioavailable iron.
Ferritin is mainly considered a cytosolic protein in many
prokaryotic and eukaryotic organisms.
However, ferritin was also found in the
nucleus, in the mitochondria and in the
endo-lysosomal compartment.
Possible functions of a secreted form of
ferritin have been also demonstrated and
Figure 2: Cell organelles
Technion – Israel Institute of Technology
Department of Biotechnology & Food Engineering
it has also been suggested that macrophages are likely the major
cellular sources for secreted serum ferritin.
Furthermore, in many immunofluorescence images published in
the literature, ferritin appears to be in an uneven distribution
which could be due to enrichment of ferritin in lysosomes. Once
in the lysosome, ferritin can be degraded and iron can be recycled.
Iron can also be stored in the lysosome. Alternatively, lysosomal
ferritin may be secreted as an additional pathway of iron export.
The current research is intended to discover the location of ferritin
in murine macrophages by immunofluorescence staining. We will
stain these cells with antibodies against ferritin and specific
organelles' proteins (simultaneously) and reveal ferritin's location
utilizing a fluorescent microscope.
Student mission / Objective:
1. Learning the basic principles of working with laboratory mice.
We have a state-of-the-art animal facility (a short tour is possible).
2. Generation of mouse bone marrow-derived macrophages.
This technique is characterized by high yield and reproducibility.
Once obtained, bone marrow-derived macrophages can be used
for a considerable number of functional and structural assays and
are commonly regarded as a model for the role of resident
macrophages in the innate immune system.
3. Acquiring knowledge on maintenance of murine primary
cultures and cell-lines (in sterile conditions). Our laboratory
comprises a cleanroom, thus general cleanroom regulations will
be thoroughly discussed.
4. Learning the principles of immunofluorescence staining method
Technion – Israel Institute of Technology
Department of Biotechnology & Food Engineering
5. Marking of ferritin and specific organelles' proteins in murine
macrophages by immunofluorescence staining and analyzing the
subcellular ferritin localization. In addition, getting familiar with
the light and the fluorescent microscopes, (present in our lab).
Figure 3: Immunofluorescence staining of ferritin (red) and lysosomal marker cathepsin D (green)
in murine macrophages (taken in our lab)
Requirements:
1. Basic background on ferritin structure, function and location
(including a thorough literature survey regarding state-of-the-art
knowledge available on this subject)
2. The willingness to work with mice (actually the students will
perform the experiments that follow dealing with mice)
3. Following the basic safety rules at the laboratory
Please read the paper (attached):
Meyron-Holtz, E. G., Moshe-Belizowski, S. & Cohen, L. A. A
possible role for secreted ferritin in tissue iron distribution. J.
Neural Transm. Vienna Austria 1996 118, 337–347 (2011)
Questions regarding the paper:
1. Why intracellular ferritin may have a role in iron trafficking?
Technion – Israel Institute of Technology
Department of Biotechnology & Food Engineering
2. What are the possible ferritin secretion pathways according to
this paper?
3. What are the possible functions of secreted ferritin in the
periphery? and in the brain?
Recommended reading material:
1. Meyron-Holtz, E. G., Moshe-Belizowski, S. & Cohen, L. A. A
possible role for secreted ferritin in tissue iron distribution. J.
Neural Transm. Vienna Austria 1996 118, 337–347 (2011).
2. Cohen, L. A. et al. Serum ferritin is derived primarily from
macrophages through a nonclassical secretory pathway. Blood
116, 1574–1584 (2010).
3. Koorts, A. M. & Viljoen, M. Ferritin and ferritin isoforms II:
protection against uncontrolled cellular proliferation, oxidative
damage and inflammatory processes. Arch Physiol Biochem 113,
55–64 (2007).
4. Blott, E. J. & Griffiths, G. M. Secretory lysosomes. Nat Rev
Mol Cell Biol 3, 122–31 (2002).
5. Oliver, Constance. “Immunomicroscopy.” In Molecular
Biomethods Handbook, edited by John M. Walker and Ralph
Rapley, 1063–79. Humana Press, 2008.
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If you have any question that might arise, feel free to contact me:
[email protected]