Dicer deletion and short RNA expression analysis in mouse embryonic stem cells
Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2008.
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| Format: | Thesis |
| Language: | eng |
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Massachusetts Institute of Technology
2008
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| Online Access: | http://hdl.handle.net/1721.1/42401 |
| _version_ | 1811086048659243008 |
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| author | Calabrese, Joseph Mauro |
| author2 | Philip A. Sharp. |
| author_facet | Philip A. Sharp. Calabrese, Joseph Mauro |
| author_sort | Calabrese, Joseph Mauro |
| collection | MIT |
| description | Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2008. |
| first_indexed | 2024-09-23T13:20:04Z |
| format | Thesis |
| id | mit-1721.1/42401 |
| institution | Massachusetts Institute of Technology |
| language | eng |
| last_indexed | 2024-09-23T13:20:04Z |
| publishDate | 2008 |
| publisher | Massachusetts Institute of Technology |
| record_format | dspace |
| spelling | mit-1721.1/424012019-04-12T11:54:41Z Dicer deletion and short RNA expression analysis in mouse embryonic stem cells Dicer deletion and short ribonucleic acid expression analysis in mouse embryonic stem cells Calabrese, Joseph Mauro Philip A. Sharp. Massachusetts Institute of Technology. Dept. of Biology. Massachusetts Institute of Technology. Dept. of Biology. Biology. Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2008. "February 2008." Vita. Includes bibliographical references. RNA interference (RNAi) manages many aspects of eukaryotic gene expression through sequence-specific interactions with RNA. Short RNAs, 20-30 nucleotides in length, guide the various effector proteins of RNAi to silence fully or partially complementary targets. The sequencing of endogenously expressed short RNA species coupled with genetic studies in various experimental organisms has revealed a role for RNAi in the silencing of protein-coding genes and repetitive elements in genomes. In mammals, it is unknown to what extent RNAi is involved in silencing processes other than the modulation of protein-coding gene expression, which is achieved through a class of short RNAs termed microRNAs (miRNAs). The work in this thesis quantitatively describes the short RNAs expressed in mouse embryonic stem (ES) cells. ES cell lines are derived from the pre-implantation blastocyst and can be cultured in vitro for extended periods while still maintaining pluripotency. It was demonstrated that approximately 130,000 5' phosphorylated short RNA molecules are present in a single ES cell. 10% of these short RNAs represent non-random fragments of larger, abundant non-coding RNA species, and have no known function. Low abundance short RNAs were discovered that cluster bidirectionally around the transcription start sites of protein-coding genes. These RNAs associate with features of active transcription, and may be evidence of widespread bidirectional initiation and pausing of RNA polymerase II in ES cells. There are on the order of 300 different miRNA species expressed in ES cells, comprising 85% of the total pool of 130,000 5' phosphorylated short RNAs. Based on experiments correlating miRNA abundance to target repression, only about 30 of these miRNAs are expected to carry significant ES cell regulatory capacity. (cont.) ES cells lacking all miRNAs do not significantly change their morphology or gene expression patterns, but do show a significant drop in growth rate compared to controls, suggesting that a major function of ES cell miRNAs may be to govern cell division. A detailed comparison of short RNAs expressed in ES cells with and without the ribonuclease Dicer strongly suggests that miRNAs are the sole regulatory molecules that function through the RNAi pathway in ES cells. Considering previous work showing that repeating elements are frequently under Dicer-dependent repression, this observation raises the possibility that mammalian miRNAs may in certain contexts function to silence repeating genomic elements in addition to protein-coding genes. by Joseph Mauro Calabrese. Ph.D. 2008-09-03T15:33:12Z 2008-09-03T15:33:12Z 2008 Thesis http://hdl.handle.net/1721.1/42401 237110765 eng M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission. http://dspace.mit.edu/handle/1721.1/7582 216 leaves application/pdf Massachusetts Institute of Technology |
| spellingShingle | Biology. Calabrese, Joseph Mauro Dicer deletion and short RNA expression analysis in mouse embryonic stem cells |
| title | Dicer deletion and short RNA expression analysis in mouse embryonic stem cells |
| title_full | Dicer deletion and short RNA expression analysis in mouse embryonic stem cells |
| title_fullStr | Dicer deletion and short RNA expression analysis in mouse embryonic stem cells |
| title_full_unstemmed | Dicer deletion and short RNA expression analysis in mouse embryonic stem cells |
| title_short | Dicer deletion and short RNA expression analysis in mouse embryonic stem cells |
| title_sort | dicer deletion and short rna expression analysis in mouse embryonic stem cells |
| topic | Biology. |
| url | http://hdl.handle.net/1721.1/42401 |
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