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- Title
- Deamplification of supernumerary centrosomes by centrosomal clustering.
- Creator
- Thomas, Ezekiel., Harriet L. Wilkes Honors College
- Abstract/Description
-
Supernumerary centrosomes can arise in a cell through a variety of methods. The presence of supernumerary centrosomes has been observed in nearly all types of cancer and promotes chromosomal instability, with rates of incident increasing as the cancer progresses. An oral squamous cell carcinoma line was treated with hydroxyurea to induce supernumerary centrosomes in the cells. NuMA was then knocked down using shRNA to promote centrosomal clustering and bipolar mitotic division in cells with...
Show moreSupernumerary centrosomes can arise in a cell through a variety of methods. The presence of supernumerary centrosomes has been observed in nearly all types of cancer and promotes chromosomal instability, with rates of incident increasing as the cancer progresses. An oral squamous cell carcinoma line was treated with hydroxyurea to induce supernumerary centrosomes in the cells. NuMA was then knocked down using shRNA to promote centrosomal clustering and bipolar mitotic division in cells with supernumerary centrosomes. Immunofluorescence with an antibody against SAS 6 accuately stained the centrioles for observation. The cells exhibiting supernumerary centrosomes undergoing bipolar mitotic division were studied to look for a possible pattern in centrosomal clustering where the majority of centrosomes are at one pole with a single centrosome at the other pole. Initial results suggest the presence of such a mechanism, which would describe a previously unknown mechanism for cells to deamplify supernumerary centrosomes by centrosomal clustering.
Show less - Date Issued
- 2012
- PURL
- http://purl.flvc.org/FAU/3359328
- Subject Headings
- Centrosomes, Cell division, Cellular signal transduction, Cancer, Genetic aspects
- Format
- Document (PDF)
- Title
- Contributions of dynactin's p150[Glued] subunit's binding domains to microtubule anchoring at the centrosome.
- Creator
- Schneider, Rebecca., Harriet L. Wilkes Honors College
- Abstract/Description
-
Intracellular transport carries out very important roles within the cell including mitosis, organization, and organelle function. In order for effective transport using the motor protein dynein, a cofactor named dynactin is required. Of dynactin's many subunits, p150[Glued] holds the most responsibility for effective microtubule organization throughout the cell and the necessary anchoring at the centrosome. P150[Glued] holds two areas of high binding potential, the CAP-Gly region and the...
Show moreIntracellular transport carries out very important roles within the cell including mitosis, organization, and organelle function. In order for effective transport using the motor protein dynein, a cofactor named dynactin is required. Of dynactin's many subunits, p150[Glued] holds the most responsibility for effective microtubule organization throughout the cell and the necessary anchoring at the centrosome. P150[Glued] holds two areas of high binding potential, the CAP-Gly region and the Basic region. Each of these binding domains have different binding potentials and affinities for microtubules. The CAP-Gly region binds tightly the microtubules for a longer period of tiem ; the Basic region binds loosely to microtubules. Throughout the course of my research, I manipulated these two regions binding affinity for microtubules and evaluated the resulting cells ability to effectively organize microtubules and anchor them properly at the centrosome.
Show less - Date Issued
- 2012
- PURL
- http://purl.flvc.org/FAU/3359326
- Subject Headings
- Cytoskeletal proteins, Microtubules, Centrosomes, Cell organelles, Formation, Molecular biology, Tubulins
- Format
- Document (PDF)
- Title
- Devising a noncancerous model system to study multipolar spindle formation.
- Creator
- Nagarsheth, Nisha., Harriet L. Wilkes Honors College
- Abstract/Description
-
Aneuploid tumor cells have characteristically unstable genomes which can be caused by mitotic defects such as multipolar spindles. Multipolarity relies upon the presence of extra centrosomes to form. However, some cells, both cancerous and noncancerous are able to avoid the formation of multipolar spindles through centrosomal clustering. Previous research has shown that there are a large number of genes whose activity contributes to the clustering activity, making analysis of individual...
Show moreAneuploid tumor cells have characteristically unstable genomes which can be caused by mitotic defects such as multipolar spindles. Multipolarity relies upon the presence of extra centrosomes to form. However, some cells, both cancerous and noncancerous are able to avoid the formation of multipolar spindles through centrosomal clustering. Previous research has shown that there are a large number of genes whose activity contributes to the clustering activity, making analysis of individual components of the process difficult. In order to better understand centrosomal clustering in cancer cells, we induced supernumerary centrosomes in a genomically normal cell line, RPE, to observe how the normal cells cope with extra centrosomes. Using colcemid to induce extra centrosomes in the RPE cell line, we observed an intact clustering mechanism in fixed cells. Further manipulation of the cells has allowed us to induce multipolarity in this cell line using various disrupters of cell-cycle checkpoint and dynein function.
Show less - Date Issued
- 2010
- PURL
- http://purl.flvc.org/FAU/3335107
- Subject Headings
- Centrosomes, Research, Cancer, Genetic aspects, Cellular signal transduction, Cell division
- Format
- Document (PDF)
- Title
- Manipulation of normal cells to produce a cancer-like mitotic phenotype.
- Creator
- Luffman, Christina., Harriet L. Wilkes Honors College
- Abstract/Description
-
Most tumors contain multiple karyotypes due to genomic instability gained through chromosomal segregational defects. The variability of genomic changes within a population makes it difficult to study specific processes without the existence of confounding mutations. My project is to create a model system for observation of mitotic defects, specifically multipolar spindles, in a normal cell line, where the genome is intact. Induction of centrosome amplification is required for formation of...
Show moreMost tumors contain multiple karyotypes due to genomic instability gained through chromosomal segregational defects. The variability of genomic changes within a population makes it difficult to study specific processes without the existence of confounding mutations. My project is to create a model system for observation of mitotic defects, specifically multipolar spindles, in a normal cell line, where the genome is intact. Induction of centrosome amplification is required for formation of multipolar spindles. Treatments with colcemid showed a 10% increase in abnormal centrosome numbers over control. However, treatment with hydroxyurea and transfection of hMPSl showed little increase. Extra centrosomes are insufficient to drive multipolarity, therefore, I am using siRNA-mediated knockdown of Nek2 or HSET to decluster the extra centrosomes. Successful declustering will preferably show an increase in multipolar frequency, allowing us to study the formation and resolution of these structres to better understand how they contribute to aneuploidy and tumor progression.
Show less - Date Issued
- 2009
- PURL
- http://purl.flvc.org/FAU/3325079
- Subject Headings
- Cell division, Karyokinesis, Cancer, Genetic aspects, Genomics, Cellular signal transduction, Centrosomes
- Format
- Document (PDF)
- Title
- Differential functionality of microtubule binding motifs in the p150[Glued] subunit of dynactin.
- Creator
- Mallen, Michael Luis., Harriet L. Wilkes Honors College
- Abstract/Description
-
Dynactin is a multi-subunit protein complex that is the required cofactor for the cytoplasmic motor protein, dynein. Dynactin is critical for cytoplasmic activities such as vesicle transport, cytoskeletal organization, membrane organization and mitotic progression as it serves to increase processivity of dynein. The main subunit in focus for this study is the sidearm extension p150Glued. On the exposed end (N-terminus) lie two different microtubule binding motifs: CAP-Gly and Basic. CAP-Gly...
Show moreDynactin is a multi-subunit protein complex that is the required cofactor for the cytoplasmic motor protein, dynein. Dynactin is critical for cytoplasmic activities such as vesicle transport, cytoskeletal organization, membrane organization and mitotic progression as it serves to increase processivity of dynein. The main subunit in focus for this study is the sidearm extension p150Glued. On the exposed end (N-terminus) lie two different microtubule binding motifs: CAP-Gly and Basic. CAP-Gly has a high binding affinity for microtubules, while Basic binds the microtubules quite weakly. It is at this location that dynactin is proposed to assist in the processivity of transporting cargo long distances in the cell, by providing additional support for the dynein molecule as it progresses along the microtubule. Analysis of the effects of overexpression of these motifs, alone or in tandem, suggested that there was no disruption of the dynein/dynactin interaction, but its ability to act in a processive manner had been perturbed. Effects on non-dynamic processes, such as Golgi localization and microtubule organization were generally weak, but effects were greater on y-tubulin organization. While this could suggest a defect in centrosome organization, we propose that it instead shows that cargo motility in the affected cells is defective, and that transport of centrosomal proteins to the centrosome is interrupted, suggesting a broader cargo-transport defect may be evident in these cells.
Show less - Date Issued
- 2007
- PURL
- http://purl.flvc.org/FAU/11613
- Subject Headings
- Cytoskeletal proteins, Tubulins, Microtubules, Centrosomes, Cell organelles, Formation
- Format
- Document (PDF)
