Download TDP-43 and FUS/TLS

TDP-43 and FUS/TLS: Emerging
Roles in RNA Processing and
Neurodegeneration
By Clotilde Lagier-Tourenne, Magdalini Polymenidou,
and Don W. Cleveland
Powerpoint Presentation by Jonah Cader
Overview:
• Amyotrophic lateral sclerosis,
commonly known as ALS or Lou
Gehrig’s disease, is an adult onset,
ultimately fatal, neurodegenerative
disorder. Over a typical course of 1-5
years, it is characterized by
progressive paralysis and the loss
of both both upper and lower
motor neurons. No cure and no
substantial clinical treatments exist.
•For years, ALS research was nearly
impossible (as over 90% of cases are
sporadic), but the identification of
toxic mutant SOD1 in ALS
pathogenesis led the advent of
molecular and genetic lines of
inquiry.
Lower Motor Neurons: Primary motor
neurons of the spinal cord and brainstemthat directly innervate skeletal muscle
Upper Motor Neuron: Originating in
higher regions of the brain (ex. the motor
cortex), they synapse on the lower motor
neurons to convey descending commands
for movement
•Since then the field has completely shifted
directions. Discoveries have been rapid as
the focus has become understanding the
SOD1 Mouse Model: Breakthrough in the
role of RNA processing/regulation in
field; genome contains a mutation in a
neurodegeneration.
gene coding for superoxide dismutase 1
(an enzyme that destroys free radicals).
•
Over 150 mutations across 3 SOD1 genes
•Most notably Cytoplasmic mislocalized
have been linked to familial ALS in
aggregates of both FUS/TLS and TDP-43
humans
have been linked to not only ALS but
numerous other neurodegenerartive
FTLD: Frontotemporal lobar degeneration –
disorders.
another neurodegenerative disorder that
causes various language and behavioral
issues.
Cytoplasmic Mislocalized Aggregates:
Also called “inclusions,” or “preinclusions,” they are abnormal protein
deposits within the cytoplasm
The Players: TDP-43 and
FUS/TLS
TDP-43:
FUS/TLS:
•Name stands for 43 kDa transactive
response (TAR) DNA-Binding protein
•Name stands for fused in
sarcoma/translocated in liposarcoma
•Protein composed of 414 amino acids,
coded for by six exons and two RNA
recognition motifs
•Also a RNA/DNA binding protein, it is
composed of 526 amino acids encoded by 15
exons
•C-terminal glycine-rich region is key to
interaction with other proteins
•Binds single-stranded RNA and doublestranded DNA. Affinity for 3’ UTR of actinstabilizing protein mRNA
•Both are multifunctional proteins, mainly localized within the nucleus (though
low levels accumulate in the cytoplasm of most cells). They play a series of
critical roles in multiple steps of RNA processing: Transcription, splicing,
transport, translation, and several other pathways
Proposed Physiological Roles of TDP-43 and
FUS/TLS
In General…
•Transcription: TDP-43 Binds single-stranded segments of promoter region (in TGrich segments), blocking downstream gene transcription. FUS/TLS assumed to
participate in general transcription (associated with a variety of machinery). DNA
damage triggers the recruitment of FUS/TLS to promoter regions of cyclin D1 by
ncRNAs, repressing CCND1
•Splicing: TDP-43 binds intronic UG segments located before alternately spliced
exons thereby increasing the likelihood of their exclusion. FUS/TLS is a component
of the spliceosome
•miRNA Processing: Identified in a Drosha-containing complex
•Nucleo-Cytoplasmic Shuttling: Both proteins move between these two regions
(roles yet unknown)
•Stress Granules: Form complexes with mRNA and other RNA binding proteins
• RNA Transport and Local Translocation: involved in moving mRNA to dendritic
spine/axonal regions where they may facilitate some local translation
Some Specifics…
•Transcription: TDP-43 associates with euchromatin (during transcription of genes).
FUS/TLS interacts with RNA polymerase II possibly influencing initiation
•Splicing: Splicing alterations and errors have been reported in sporadic cases. The
roles of TDP-43 and FUS/TLS in these errors are still being investigated
•miRNA Processing: Associate with Drosha, the protein that regulates the first step
of miRNA maturation. TDP-43 may be involved in Dicer complex-mediated cleaving
•RNA Regulation, Localization, Translation, and decay: Critical components of stress
granules (cytoplasmic foci made of mRNA and other complexes that delay translation
under stress conditions). TDP-43 may help stabilize and transport low molecular
weight neurofilament mRNA to injuries sites. (NFL protein necessary for axonal
repair)
•Maintenance of Genome Stability: TDP-43 is necessary for early mouse
embryogenesis (knockdown of TARDBP gene is embryonically lethal). FUS/TLS is very
important to genomic integrity as it helps initiate DNA repair by homologous
recombination according to several new studies
TDP-43 and FUS TLS in ALS
Pathology
•Redistribution of TDP-43 occurs early on in the progression of ALS. For some
reason, mutated TDP-43 proteins clear out of the nucleus, forming ubiquitinpositive, alpha –synuclein- and tau negative, cytoplasmic aggregates
• Similarly, abnormal FUS/TLS (primarily nuclear) cytoplasmic inclusions were found
in neuronal and glial cells of patients presenting classic ALS phenotypes. These
aggregates were were NOT immunoreceptive for TDP-43, implying that that
FUS/TLS-driven neurodegenerative pathways act independently of those related to
TDP-43 inclusions.
•Mutated TDP-43 mouse models presented a very fast, progressive paralysis (similar
to human ALS) . Moreover, overexpression of wild-type (normal) TDP-43 in primary
neurons caused cell death
•Studies of the link between morphologic changes and individual neuronal survival
supported the idea that cytoplasmic TDP-43 is toxic to rat cortical neurons
•That said, TDP-43 has been observed to undergo quick, transient cytoplasmic
translocation in response to stress. The aforementioned aggregates may in fact be
neuroprotective mechansims to cope with stress that only become toxic after the
recruitment of yet unidentified binding partners
•According to one paper, TDP-43 interruption can cause motor problems in
Drosophilia. Some of the negative effects of mutated TDP-43 metabolism may be
attributable to a loss of function .
Neuronal TDP-43 cytoplasmic
inclusions in a FTLD Patient
This is Your Research Lab…
The three prevailing hypothesis on the roles of
TDP-43 and FUST/TLS in ALS pathogenesis are:
1. Toxic gain of function: The cytoplasmic
aggregates are neurotoxic, poisoning motor
neurons and causing ALS
2. Toxic Loss of Function: The mislocalization and
mutation of these two proteins means that
several essential cellular processes are not
proceeding and/or are severely impaired. As a
result the motor neurons become diseased
3. Natural Response to Stress: The cytoplasmic
inclusions are abnormal stress granules that
form in response to the pressures ALS places
on the nervous system. (The aggregates are
natural responses to – not symptoms of – the
disease)
Based on your knowledge from this presentation and the paper itself, please
discuss and debate which line of inquiry you would pursue (as a group) in your
fictional motor neuron research lab
Final Thoughts
•Some combination of TDp-43 and/or FUS/TLS inclusions have been found in
cases of numerous other neurodegenerative disorders including Alzheimer’s,
dementia, Down syndrome, Parkinson’s, Huntington’s, and various myopathies.
(See page R50 of the article).
•While the exact pathogenic roles of the abnormal forms of FUS/TLS and TDP-43
have yet to be discovered, RNA/DNA binding protein research has the potential
to significantly affect not only ALS but also the study of neurodegenerative
diseases as a whole. (And, of course, the end goal of developing cures).
Sources
Information comes from the journal article itself and photos were found at the
following URLs:
http://sandiegocaregiversblog.com/wp-content/uploads/2010/06/Lou_Gehrig.jpg
http://www.nature.com/nature/journal/v445/n7125/images/445242a-i2.0.jpg
http://www.springerimages.com/Images/Biomedicine/1-10.1007_s00429-0100263-z-3