Investigating the role of a 25 kDa truncated isoform of estrogen receptor alpha (ERα) in breast cancer
(2024) MOBM02 20241Degree Projects in Molecular Biology
- Abstract
- Background: Estrogen receptor alpha (ERα) is a protein that belongs to the nuclear receptor family. It is regulating the expression of genes involved in cell cycle progression, proliferation, and apoptosis. Approximately 75% of breast cancers express ERα, making it an important target for treatment. Researchers have identified various isoforms of ERα, with the most well-characterized one being a 66 kDa protein. In a recent study from our group, an alternative splicing event was identified. The resulting protein is a truncated form of approximately 25 kDa. Preliminary data suggests that knockdown of the truncated protein leads to increased sensitivity to fulvestrant. The aim of this study is to understand the function of this truncated... (More)
- Background: Estrogen receptor alpha (ERα) is a protein that belongs to the nuclear receptor family. It is regulating the expression of genes involved in cell cycle progression, proliferation, and apoptosis. Approximately 75% of breast cancers express ERα, making it an important target for treatment. Researchers have identified various isoforms of ERα, with the most well-characterized one being a 66 kDa protein. In a recent study from our group, an alternative splicing event was identified. The resulting protein is a truncated form of approximately 25 kDa. Preliminary data suggests that knockdown of the truncated protein leads to increased sensitivity to fulvestrant. The aim of this study is to understand the function of this truncated isoform.
Methods: We conducted a functional analysis of the truncated isoform in three different cell lines - HCC1428-FR, MCF7, and HepG2. We employed several techniques such as transient transfection, western blotting, real-time RT-PCR, cell proliferation with alamarBlue, and luciferase reporter gene assay.
Results: Our research findings indicate knockdown of the truncated isoform led to a decrease in full-length ERα on the protein level, while knocking down the full-length ERα resulted in an upregulation of the truncated isoform. Overexpression of the truncated isoform significantly upregulated the full-length ERα, indicating a possible correlation between the two isoforms. However, the role of the truncated isoform in response to fulvestrant and the results from subcellular localization and luciferase assays were inconclusive.
Conclusion: Although we have observed some promising results, it is important to carefully evaluate the conditions under which the previous experiments were conducted and optimize the protocols accordingly. Our future experiments involve repeating the luciferase assays in different cell lines, blocking the full-length isoform with 4-hydroxytamoxifen or using a higher concentration of fulvestrant, immunoprecipitate the full-length protein with a C-terminal antibody under non-denaturing conditions and mutate sites susceptible to SUMOylation. (Less) - Popular Abstract
- Breast cancer: Unraveling the role of a novel estrogen receptor isoform
Female breast cancer has become the most common cancer diagnosed worldwide, and its incidence is expected to rise significantly in the coming decades. One of the most important biomarkers in the clinic is estrogen receptor (ER). ER is a protein that binds with estrogen, a hormone that plays a role in many body functions, including breast development and menstruation. When estrogen binds to ER, it activates the protein, which then sends signals to the cell. In breast cells, ER plays a role in how cells grow and develop. Our study focuses on the role of the ER and how variations of it affect breast tumors.
ER is present in about 7 out of 10 breast cancers, and it... (More) - Breast cancer: Unraveling the role of a novel estrogen receptor isoform
Female breast cancer has become the most common cancer diagnosed worldwide, and its incidence is expected to rise significantly in the coming decades. One of the most important biomarkers in the clinic is estrogen receptor (ER). ER is a protein that binds with estrogen, a hormone that plays a role in many body functions, including breast development and menstruation. When estrogen binds to ER, it activates the protein, which then sends signals to the cell. In breast cells, ER plays a role in how cells grow and develop. Our study focuses on the role of the ER and how variations of it affect breast tumors.
ER is present in about 7 out of 10 breast cancers, and it is associated with a more favorable prognosis. This is because ER-positive breast cancers tend to grow more slowly and are more responsive to treatment with drugs against the ER. Today we know that the ER has multiple variations, called isoforms. These isoforms can differ slightly in structure and function compared to the standard ER protein. Importantly, some isoforms might influence how breast cancer cells grow and respond to treatment.
Our research group has recently discovered a new isoform, and this project aims to study its role in breast cancer cells. We have developed experimental tools to study the biological function of this isoform in breast cancer cells grown in the laboratory. We examined the function of the isoform on protein and RNA level, we assessed its influence on the standard ER, we evaluated its effect on cell growth in combination with a drug called fulvestrant, and we investigated its localization within the cell. Our results show that when the standard ER is reduced, this isoform becomes more abundant. This may indicate that the cells use this isoform to compensate for the lack of the standard receptor. Additionally, we suspect that this isoform affects the effectiveness of fulvestrant, but further experiments are needed to understand the biological mechanism.
Understanding how the isoforms affect the response to treatments can help physicians to choose the most appropriate treatment. This project highlights the potential role of ER isoforms influencing breast cancer development and progression.
Master’s Degree Project in Molecular Biology 30 credits 2024 Department of Biology, Lund University
Advisors: Helena Persson, Juliane Albrecht
Faculty of Medicine, Department of Clinical Sciences Lund, Oncology, Lund University, Sweden (Less)
Please use this url to cite or link to this publication:
http://lup.lub.lu.se/student-papers/record/9176232
- author
- Prifti, Izabela
- supervisor
- organization
- course
- MOBM02 20241
- year
- 2024
- type
- H2 - Master's Degree (Two Years)
- subject
- language
- English
- id
- 9176232
- date added to LUP
- 2024-10-07 15:55:39
- date last changed
- 2024-10-07 15:55:39
@misc{9176232,
abstract = {{Background: Estrogen receptor alpha (ERα) is a protein that belongs to the nuclear receptor family. It is regulating the expression of genes involved in cell cycle progression, proliferation, and apoptosis. Approximately 75% of breast cancers express ERα, making it an important target for treatment. Researchers have identified various isoforms of ERα, with the most well-characterized one being a 66 kDa protein. In a recent study from our group, an alternative splicing event was identified. The resulting protein is a truncated form of approximately 25 kDa. Preliminary data suggests that knockdown of the truncated protein leads to increased sensitivity to fulvestrant. The aim of this study is to understand the function of this truncated isoform.
Methods: We conducted a functional analysis of the truncated isoform in three different cell lines - HCC1428-FR, MCF7, and HepG2. We employed several techniques such as transient transfection, western blotting, real-time RT-PCR, cell proliferation with alamarBlue, and luciferase reporter gene assay.
Results: Our research findings indicate knockdown of the truncated isoform led to a decrease in full-length ERα on the protein level, while knocking down the full-length ERα resulted in an upregulation of the truncated isoform. Overexpression of the truncated isoform significantly upregulated the full-length ERα, indicating a possible correlation between the two isoforms. However, the role of the truncated isoform in response to fulvestrant and the results from subcellular localization and luciferase assays were inconclusive.
Conclusion: Although we have observed some promising results, it is important to carefully evaluate the conditions under which the previous experiments were conducted and optimize the protocols accordingly. Our future experiments involve repeating the luciferase assays in different cell lines, blocking the full-length isoform with 4-hydroxytamoxifen or using a higher concentration of fulvestrant, immunoprecipitate the full-length protein with a C-terminal antibody under non-denaturing conditions and mutate sites susceptible to SUMOylation.}},
author = {{Prifti, Izabela}},
language = {{eng}},
note = {{Student Paper}},
title = {{Investigating the role of a 25 kDa truncated isoform of estrogen receptor alpha (ERα) in breast cancer}},
year = {{2024}},
}