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Mechanistic Analysis of The Function of A Long Non-coding RNA

ZHANG, BING
http://rave.ohiolink.edu/etdc/view?acc_num=case1427212306

Abstract Details

2015, Doctor of Philosophy, Case Western Reserve University, Biochemistry.
The ubiquitous presence of long non-coding RNAs (lncRNAs) in higher eukaryotes points to the importance of understanding the mechanism of their functions. Currently, a large number of studies on the mechanism of lncRNAs’ functions have been focused on identifying the correlative relationships between lncRNAs and a variety of cellular processes and diseases. In contrast, very few cases of studies on the detailed molecular mechanism of lncRNAs’ functions have been reported. To gain insight into the mechanistic actions of lncRNAs, we selected an example (lncRNA BORG) from this lncRNA group and performed detailed a mechanistic analysis on its functions. BORG is an intergenic lncRNA that was originally discovered as a target gene of the Bone Morphogenetic Protein (BMP) pathway. In order to gain insight into the mechanism of action of nuclear lncRNA BORG, we first analyzed the sequence requirements for its nuclear localization. Our study showed that the subcellular localization of BORG was not dependent on the context or level of its expression or decay, but rather depended on the primary sequence of the mature, spliced transcript. Mutational analyses indicated that nuclear localization of BORG was mediated through a novel RNA motif. Interestingly, the presence of this motif in other cellular RNAs showed a direct correlation with nuclear localization. After that, we explored the mechanism of action of BORG in cellular stress response. The expression of BORG can be induced by stressful stimuli in several neuronal and non-neuronal cell types, including cells of mesodermal and ectodermal origin. After exposure to stress, BORG knockdown cells exhibited an impaired cellular stress response and a much lower survival rate compared to controls, while the BORG overexpression cells showed an elevated level of stress response and a cellular stress-resistant phenotype. Analysis of the stress response factors indicated that some, but not all, heat shock proteins had a higher basal level in the overexpression cells even in the absence of stress and thus, the elevated expression of the lncRNA seemed to “pre-condition” the cells for tolerance to stress. Interestingly, in the BORG knockdown cells, the induction of heat shock proteins after stress was both delayed and reduced in magnitude. RNA pull-down assay and mass spectrometry showed that BORG is associated with the Replication Protein A (RPA) complex in vivo. Knockdown of RPA1 or HSF1 abolished the protective effect of BORG overexpression and blocked the induction of heat shock proteins. These data indicate that BORG plays an important role in regulation of the stress response and cellular survival under stressful conditions, likely through modulation of the function of RPA complex and HSF1. In summary, our study has identified a novel RNA nuclear localization motif and clarified the function of lncRNA BORG in the cellular stress response. These findings have demonstrated how the sequence of RNAs mediates their function and how lncRNAs can play critical roles in regulating fundamental cellular processes in higher eukaryotic organisms.
SABA VALADKHAN (Advisor)
165 p.
Biochemistry
lncRNA

Recommended Citations

Citations

  • ZHANG, B. (2015). Mechanistic Analysis of The Function of A Long Non-coding RNA [Doctoral dissertation, Case Western Reserve University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=case1427212306

    APA Style (7th edition)

  • ZHANG, BING. Mechanistic Analysis of The Function of A Long Non-coding RNA. 2015. Case Western Reserve University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=case1427212306.

    MLA Style (8th edition)

  • ZHANG, BING. "Mechanistic Analysis of The Function of A Long Non-coding RNA." Doctoral dissertation, Case Western Reserve University, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=case1427212306

    Chicago Manual of Style (17th edition)

case1427212306
315
© 2015, all rights reserved.
This open access ETD is published by Case Western Reserve University School of Graduate Studies and OhioLINK.