Abstract
Sarcomeres, highly ordered assemblages of hundreds of proteins,
perform the work of
muscle contraction. Despite increasing knowledge of sarcomere
components and their
functions, we still do not have a clear understanding about how
sarcomeres are assembled
and maintained during muscle contraction. Our laboratory studies
sarcomere assembly in
the model genetic organism C. elegans, focusing on the
function of the giant muscle
protein UNC-89. To understand the molecular mechanisms by which
UNC-89 is
assembled at the M-line, and how UNC-89 performs its functions, our
laboratory is
identifying its binding partners, and studying their
functions.
My thesis focused on finding binding partners for two regions of
UNC-89: the C-
terminal protein kinase region, and the N-terminal Ig domains 1-5.
I discovered that the
PK1 protein kinase domain and interkinase region interact with
LIM-9 (FHL in humans),
and that LIM-9 interacts with SCPL-1, a CTD type phosphatase,
previously identified by
the lab as interacting with the kinase domains of UNC-89. I propose
two structural
models for the function of these interactions at the M-line.
I have discovered a new binding partner for the N-terminal region
of UNC-89, called
CPNA-1. CPNA-1 contains a conserved "copine domain" with weak
homology to the
extracellular portions of integrins, and largely unknown function.
I have defined a new
category of copine domain containing proteins, the "atypical" ones.
CPNA-1 specific
antibodies localize to integrin adhesion complexes (M-lines and
dense bodies) of
nematode body wall muscle. I found that CPNA-1 binds to the M-line
proteins UNC-89,
LIM-9 (FHL), SCPL-1, UNC-96, and a protein common to the M-line and
dense body,
PAT-6 (actopaxin). A genomic deletion for cpna-1, gk266,
displays the typical Pat
(Paralyzed arrested at two-fold) phenotype. By localizing
previously characterized
muscle adhesion complex proteins in cpna-1 mutant embryos,
and localizing CPNA-1 in
other Pat mutants, I have placed CPNA-1 in the M-line/dense body
assembly pathway of
embryonic muscle. By using RNAi and mutants, I have begun to define
a role for CPNA-
1 in adult muscle. I conclude with a model that PAT-6 recruits
CPNA-1, and in turn,
CPNA-1 recruits additional proteins (UNC-89, LIM-9, SCPL-1, UNC-96)
to the M-line.
Table of Contents
Table of Contents
1. Chapter 1:
Introduction
1.1 Muscle
Organization
1.2 Use of C.
elegans to Study Muscle Sarcomere Organization and
Assembly
1.3 Giant Polypeptides in
Muscle
1.3.1 C.elegans
Giants
1.3.2 Human
Titin
1.3.3 Obscurin (Human
UNC-89)
1.3.4
UNC-89
2. Chapter 2: A LIM-9
(FHL) / SCPL-1 (SCP) Complex Interacts with the C-terminal Protein
Kinase Regions of UNC-89 (Obscurin) in C. elegans
Muscle
2.1
Introduction
2.2
Results
2.3
Discussion
2.4 Materials and
Methods
3. Chapter 3: CPNA-1, A
Novel Copine Containing Protein, Links Integrin Associated Protein
PAT-6 (Actopaxin) to the Giant Protein UNC-89 (Obscurin) at Muscle
Adhesion Site in C. elegans
3.1
Introduction
3.2
Results
3.3
Discussion
3.4 Materials and
Methods
4. Chapter 4: Summary and
Future Directions
4.1
CPNA-1
4.2 A LIM-9/SCPL-1
Complex Interacts with the C-terminal Portion of
UNC-89
5.
References
About this Dissertation
Rights statement
- Permission granted by the author to include this thesis or dissertation in this repository. All rights reserved by the author. Please contact the author for information regarding the reproduction and use of this thesis or dissertation.
| School |
|
|---|
| Department |
|
|---|
| Subfield / Discipline |
|
|---|
| Degree |
|
|---|
| Submission |
|
|---|
| Language |
|
|---|
| Research Field |
|
|---|
| Keyword |
|
|---|
| Committee Chair / Thesis Advisor |
|
|---|
| Committee Members |
|
|---|