Skip to Main Content
Frequently Asked Questions
Submit an ETD
Global Search Box
Need Help?
Keyword Search
Participating Institutions
Advanced Search
School Logo
Files
File List
SLai_Thesis_final.pdf (12.55 MB)
ETD Abstract Container
Abstract Header
Defective tRNA Processing by RNase P Contributes to Neurodegeneration in Mice
Author Info
Lai, Stella Myra
ORCID® Identifier
http://orcid.org/0000-0001-7107-0629
Permalink:
http://rave.ohiolink.edu/etdc/view?acc_num=osu1493997936911358
Abstract Details
Year and Degree
2017, Master of Science, Ohio State University, Anatomy.
Abstract
Ribonuclease P (RNase P) is an essential enzyme that catalyzes the 5' maturation of transfer RNAs (tRNAs), which function as adaptor molecules during the synthesis of proteins from messenger RNAs. The presence of two concurrent mutations in C57BL6/J (B6J)-
nmf205
-/-
mice induced progressive retinal degeneration and neuronal death: a phenotypically silent, single-nucleotide substitution (C50U) in
n-Tr20
, a central nervous system-specific, nuclear-encoded tRNA
Arg
UCU
isodecoder; and a loss-of-function mutation in guanosine triphosphate binding protein 2 (GTPBP2), a ribosome-release factor involved in the recycling of stalled translation machinery [Ishimura et al. (2014) Science,
345
: 455-459]. High levels of immature
n-Tr20-C50U
, coupled with markedly low amounts of mature
n-Tr20-C50U
, implicate defective processing of mutant
n-Tr20
by RNase P as one of the underlying causes of neurodegeneration in B6J-
nmf205
-/-
mice. Thus, the objective of this study was to uncover the molecular basis for this processing defect and determine whether it arises from either (1) spatiotemporal variations in the protein subunit composition of brain RNase P that render the enzyme less permissive for cleavage of mutant substrates or (2) alterations in the secondary and tertiary structure of the mutant tRNA that make it a poor substrate for RNase P cleavage. RNase P was isolated from mouse and human brain and liver tissues for the first time, and these tissue-specific isoforms were characterized to uncover any differences in their ability to process
n-Tr20-C50U
. Surprisingly, under near-physiological conditions (150 mM NaCl and 1 mM MgCl
2
), all four RNase P enzymes were defective in processing the mutant substrate, and increasing the Mg
2+
concentration to 5 mM only partially rescued the defect. These findings suggest that an altered structure must preclude robust processing of
n-Tr20-C50U
by RNase P. Subsequent thermal denaturation experiments to study the effect of the single-nucleotide substitution on the stability of
n-Tr20
confirmed this notion. However, contrary to the numerous reported examples of destabilizing tRNA mutations, the C50U mutation
stabilized
the structure of the tRNA dramatically, increasing the
T
m
of the mutant by 5-10°C relative to that of the wild-type tRNA. This increase in melting temperature likely reflects significant changes in the secondary and tertiary structure of the tRNA. High-resolution crystallographic studies are needed to deduce the structural impact of the C50U mutation. More broadly, gaining structural insights into the conformational changes wrought by individual tRNA mutations will help further our understanding of the roles they play in causing disease.
Committee
Stephen Kolb (Advisor)
Laura Boucher (Advisor)
Pages
141 p.
Subject Headings
Anatomy and Physiology
;
Biochemistry
;
Molecular Biology
Keywords
RNase P
;
tRNA processing
;
neurodegeneration
Recommended Citations
Refworks
EndNote
RIS
Mendeley
Citations
Lai, S. M. (2017).
Defective tRNA Processing by RNase P Contributes to Neurodegeneration in Mice
[Master's thesis, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1493997936911358
APA Style (7th edition)
Lai, Stella.
Defective tRNA Processing by RNase P Contributes to Neurodegeneration in Mice.
2017. Ohio State University, Master's thesis.
OhioLINK Electronic Theses and Dissertations Center
, http://rave.ohiolink.edu/etdc/view?acc_num=osu1493997936911358.
MLA Style (8th edition)
Lai, Stella. "Defective tRNA Processing by RNase P Contributes to Neurodegeneration in Mice." Master's thesis, Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu1493997936911358
Chicago Manual of Style (17th edition)
Abstract Footer
Document number:
osu1493997936911358
Download Count:
39
Copyright Info
© 2017, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.