Muscleblind Like Splicing Regulator 1 (MBNL1) Associates with Endosomes in Mouse Neuroblastoma Cells Open Access

Abstract

Myotonic Dystrophy Type 1 (DM1) is a genetic disorder that affects multiple organ systems, including the central nervous system, and causes symptoms such as cognitive difficulties and hypersomnia. Muscleblind-like (MBNL) proteins are a family of RNA-binding proteins (RBPs) that contain zinc-finger domains. In DM1, MBNL proteins are sequestered in the nucleus by CUG trinucleotide repeats. It is unclear how the sequestration of MBNLs contributes to DM1. RBPs participate in RNA localization, associating with motor proteins and/or endomembranes during transport. Recent findings have shown that RNA is able to be transported with motile endosomes, leading to the possibility that Muscleblind Like Splicing Regulator 1 (MBNL1) transports mRNA by interacting with the endomembrane system. This thesis investigates MBNL1 trafficking within the endomembrane system. I hypothesized that MBNL1 associates with varying degrees with different kinds of endosomes. Here, we show that MBNL1 colocalized with early, late, recycling endosomes, and exosomes/lysosomes. Using Rab5, Neep21, and EEA1 antibodies that correspond to early endosomes, I demonstrated that MBNL1 colocalized to a greater degree to early endosomes compared to late endosomes, recycling endosomes, and lysosomes. From these findings, I explored how Rab5 function affects MBNL1 mobility and trafficking in the endomembrane system. I showed that MBNL1 exhibits a trend towards more

colocalization with more in constitutively active Rab5 than wild-type Rab5. Live imaging showed a trend towards greater MBNL1 movement in both constitutively active Rab5 and dominant negative Rab5 as compared to wild-type Rab5. Results show endosomes, especially early endosomes, are involved in MBNL1 trafficking and anchoring. This thesis motivates further research using live cell imaging to study if the mobility of MBNL1 is disturbed when early endosome mobility is disturbed. More research is needed to understand the underlying mechanisms of MBNL1 transportation and anchoring to regulate mRNA localization and translation.

Table of Contents

Introduction .......................................................................... 1 Methods ............................................................................... 12 Results ................................................................................. 16 Discussion ............................................................................ 19 Figures ................................................................................ 23 References ........................................................................... 29

About this Honors Thesis

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School	Emory College
Department	Chemistry
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Biology, Molecular Chemistry, Biochemistry
Keyword	MBNL endosomes myotonic dystrophy
Committee Chair / Thesis Advisor	Gary Bassell, Emory University
Committee Members	Khalid Salaita, Emory University Vincent Conticello, Emory University

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