Type

Text

Type

Dissertation

Advisor

Czaplinski, Kevin | Halegoua, Simon | Talmage, David | Twiss, Jeffery.

Date

2015-08-01

Keywords

Biology

Department

Department of Neuroscience.

Language

en_US

Source

This work is sponsored by the Stony Brook University Graduate School in compliance with the requirements for completion of degree.

Identifier

http://hdl.handle.net/11401/76587

Publisher

The Graduate School, Stony Brook University: Stony Brook, NY.

Format

application/pdf

Abstract

Abstract The local translation of trafficked mRNAs temporally and spatially regulates protein expression. In neurons, mRNAs are trafficked to both axons and dendrites and the local translation of these mRNAs is important for axon guidance as well as synaptic plasticity. The active trafficking of mRNAs involves the interaction between cis¬¬-acting localization elements, known as `zipcodes,' and trans-acting factors, including RNA-binding proteins. However, the exact molecular mechanism of mRNA localization in the mammalian nervous system remains unknown. To better understand this process, the zipcode element of  -actin (Actb) mRNA was used to identify putative  -actin zipcode binding proteins from the rodent brain (bZBPs). Hnrnpab was confirmed as a bZBP and was found to have an unexpected isoform dependent specificity. The larger isoform, Hnrnpab1, is specific for Actb mRNA and the zipcode localization element. The alternatively spliced isoform, Hnrnpab2, interacts with the zipcode element and the 5' UTR of Actb mRNA as well as  -actin (Actg) mRNA. Analysis using a novel fluorescent in situ hybridization method demonstrated a decrease in Actb mRNA in the periphery of cells in the absence of Hnrnpab. This effect can be rescued only with the Hnrnpab1 isoform, suggesting a distinct function in Actb mRNA localization. Mice lacking Hnrnpab show a number of changes in protein expression which suggest a role in nervous system development and glutamate signaling. Hnrnpab-/- neural stem and progenitor cells undergo altered differentiation patterns in culture, and mature Hnrnpab-/- neurons demonstrate increased sensitivity to glutamate-induced excitotoxicity. These studies represent an important step in understanding the underlying molecular mechanism of mRNA trafficking by identifying several putative localization factors using the localization element of Actb mRNA and establishing Hnrnpab1 as a zipcode binding protein, which mediates Actb mRNA localization. They also explore the role of Hnrnpab in the nervous system and provide evidence for isoform dependent functions. | Abstract The local translation of trafficked mRNAs temporally and spatially regulates protein expression. In neurons, mRNAs are trafficked to both axons and dendrites and the local translation of these mRNAs is important for axon guidance as well as synaptic plasticity. The active trafficking of mRNAs involves the interaction between cis¬¬-acting localization elements, known as `zipcodes,' and trans-acting factors, including RNA-binding proteins. However, the exact molecular mechanism of mRNA localization in the mammalian nervous system remains unknown. To better understand this process, the zipcode element of  -actin (Actb) mRNA was used to identify putative  -actin zipcode binding proteins from the rodent brain (bZBPs). Hnrnpab was confirmed as a bZBP and was found to have an unexpected isoform dependent specificity. The larger isoform, Hnrnpab1, is specific for Actb mRNA and the zipcode localization element. The alternatively spliced isoform, Hnrnpab2, interacts with the zipcode element and the 5' UTR of Actb mRNA as well as  -actin (Actg) mRNA. Analysis using a novel fluorescent in situ hybridization method demonstrated a decrease in Actb mRNA in the periphery of cells in the absence of Hnrnpab. This effect can be rescued only with the Hnrnpab1 isoform, suggesting a distinct function in Actb mRNA localization. Mice lacking Hnrnpab show a number of changes in protein expression which suggest a role in nervous system development and glutamate signaling. Hnrnpab-/- neural stem and progenitor cells undergo altered differentiation patterns in culture, and mature Hnrnpab-/- neurons demonstrate increased sensitivity to glutamate-induced excitotoxicity. These studies represent an important step in understanding the underlying molecular mechanism of mRNA trafficking by identifying several putative localization factors using the localization element of Actb mRNA and establishing Hnrnpab1 as a zipcode binding protein, which mediates Actb mRNA localization. They also explore the role of Hnrnpab in the nervous system and provide evidence for isoform dependent functions. | 135 pages

Download

Share

COinS
 
 

To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.