Presentation made by Dr. Paul Taylor on October 30, 2015 at the Alzforum-hosted live webinar titled "Fluid Business: Could “Liquid” Protein Herald Neurodegeneration?"
More information and the recording of the session available at http://www.alzforum.org/webinars/fluid-business-could-liquid-protein-herald-neurodegeneration
Presentation made by Dr. Simon Alberti on October 30, 2015 at the Alzforum-hosted live webinar titled "Fluid Business: Could “Liquid” Protein Herald Neurodegeneration?"
More information and the recording of the session available at http://www.alzforum.org/webinars/fluid-business-could-liquid-protein-herald-neurodegeneration
Regulation of KDM5 by multiple cofactors regulates cancer and stem cellsChristopher Wynder
Presentation of data regarding proteins that regulate the activity of KDM5b.
The studies use multiple disciplines including in vitro enzymology, ES cell studies of differentiation, Mass spectrometry to detect protein protein interactions.
These studies resulted in a comprehensive view of KDM5b function. It required development of at least three novel assays that are focused on moving epigenetic research from yeast and HeLa cell types to primary, clinically relevant cell types.
The techniques have been successfully used in Embryonic stem cells (human and mouse), Neural stem cells (mouse and patient derived as well as iPSCs.
USMLE Step 1 Molecular Biology and Biochemistry reviewAbril Santos
This is a review for the USMLE Step 1 exam for Molecular Biology and Biochemistry.
It contains anything you need to know for your exam in pictures, tables and diagrams.
Presentation made by Dr. Simon Alberti on October 30, 2015 at the Alzforum-hosted live webinar titled "Fluid Business: Could “Liquid” Protein Herald Neurodegeneration?"
More information and the recording of the session available at http://www.alzforum.org/webinars/fluid-business-could-liquid-protein-herald-neurodegeneration
Regulation of KDM5 by multiple cofactors regulates cancer and stem cellsChristopher Wynder
Presentation of data regarding proteins that regulate the activity of KDM5b.
The studies use multiple disciplines including in vitro enzymology, ES cell studies of differentiation, Mass spectrometry to detect protein protein interactions.
These studies resulted in a comprehensive view of KDM5b function. It required development of at least three novel assays that are focused on moving epigenetic research from yeast and HeLa cell types to primary, clinically relevant cell types.
The techniques have been successfully used in Embryonic stem cells (human and mouse), Neural stem cells (mouse and patient derived as well as iPSCs.
USMLE Step 1 Molecular Biology and Biochemistry reviewAbril Santos
This is a review for the USMLE Step 1 exam for Molecular Biology and Biochemistry.
It contains anything you need to know for your exam in pictures, tables and diagrams.
Genetic Information Transfer (Biology for Engineers)Dr. Arun Sharma
Information Transfer: Purpose: The molecular basis of coding and
decoding genetic information is universal. Molecular basis of information
transfer. DNA as a genetic material. Hierarchy of DNA structure- from
single stranded to double helix to nucleosomes. Concept of genetic code.
Universality and degeneracy of genetic code. Define gene in terms of
complementation and recombination.
Tumor suppression and inflammation: controlling the senescence associated se...adamfreund
This is the powerpoint presentation from a talk I gave at a conference in October, 2009. It will be hard to follow without the spoken part, but it will hopefully give anyone who is interested a brief introduction to my thesis research.
Genetic Information Transfer (Biology for Engineers)Dr. Arun Sharma
Information Transfer: Purpose: The molecular basis of coding and
decoding genetic information is universal. Molecular basis of information
transfer. DNA as a genetic material. Hierarchy of DNA structure- from
single stranded to double helix to nucleosomes. Concept of genetic code.
Universality and degeneracy of genetic code. Define gene in terms of
complementation and recombination.
Tumor suppression and inflammation: controlling the senescence associated se...adamfreund
This is the powerpoint presentation from a talk I gave at a conference in October, 2009. It will be hard to follow without the spoken part, but it will hopefully give anyone who is interested a brief introduction to my thesis research.
The discovery of the nuclear factor TDP-43 involvement in neurodegenerative disease has increased significantly the general interest on the characteristics of this protein. The aberrant localization and aggregation of TDP-43 in affected tissues coupled with the tight auto regulation of TDP 43 cellular levels has suggested novel pathways for neurodegeneration. TDP 43 is predominantly a nuclear protein that shuttles between nucleus and cytoplasm. In disease neurons TDP 43 mislocalize to cytoplasmic inclusions with devastating consequences on neuronal survival. These cytoplasmic aggregation disrupts the TDP-43 control of its own cellular level. In fact autoregulation is mediated byan unusual splicing event in the 3’UTR of its pre mRNA for which is essentiial the presence of TDP 43 in the nucleus. In addition animal models and highthroughput assays have recently highlighted the role played by this protein in the regulation of hundreds of nuclear and cytoplasmic RNA transcripts, many of them belonging to key genes for neuronal metabolism. A model has been developed to study the determinants of the aggregation process and the impact of the latter on neuronal function. Animal models of the disease have been developed in different species mainly mice and flies.
Genetics of Frontotemporal lobar degenerationLeena Shingavi
It describes the overview of the genetics of early onset neurodegenerative dementia with pathological involvement of the frontal and temporal lobe. A diagrammatic representation of the known mutation types and the protein domains involved in the implicated genes and/or protein function in the causation of FTLD is also depicted. The data is as updated in 2018.
Plant epigenetic memory in plant growth behavior and stress response. Sally M...CIAT
Speaker: Sally Mackenzie, Lloyd and Dottie Huck Chair for Functional Genomics, Department of Biology, Pennsylvania State University. Fellow in the American Society of Plant Biologists and the American Association for the Advancement of Science (AAAS).
Event: Robert D. Havener Seminar on “Innovations for Crop Productivity”.
http://ciat.cgiar.org/event/robert-d-havener-seminar-on-innovations-for-crop-productivity/
Moshe Szyf, PhD Presentation at 2016 Science of HOPE
The workshop will discuss basic epigenetic mechanisms and their role in setting up gene expression programs as well as review evidence from animal and human studies illustrating how epigenetic processes might be mediating the impact of the social and physical environment on the mental and physical health of children as well as adults. The implications of epigenetics for developing social policies, new interventions, diagnostics and therapeutics will be presented.
First Identification of Role TMEM106B in FTDAlzforum
Presentation made by Rosa Rademakers on the 20th of April, 2017, at the live webinar hosted by Alzforum: http://www.alzforum.org/webinars/webinar-cortex-aging-too-fast-blame-tmem106b-and-progranulin
Presentation made by Jernej Ule on the 20th of April, 2017, at the live webinar hosted by Alzforum: http://www.alzforum.org/webinars/webinar-cortex-aging-too-fast-blame-tmem106b-and-progranulin
Presentation made by Tony Wyss-Coray on the 20th of April, 2017, at the live webinar hosted by Alzforum: http://www.alzforum.org/webinars/webinar-cortex-aging-too-fast-blame-tmem106b-and-progranulin
Presentation made by Abeliovich and Rhinn on the 20th of April, 2017, at the live webinar hosted by Alzforum: http://www.alzforum.org/webinars/webinar-cortex-aging-too-fast-blame-tmem106b-and-progranulin
Marc Dhenain Alzforum Webinar - Dec 7, 2016Alzforum
Presentation made at the Alzforum's live webinar of December 5, 2016, titled "Is Alzheimer’s Disease a Uniquely Human Disorder?" - review additional information and recording at www.alzforum.org/
Peter Nelson Alzforum Webinar - Dec 7, 2016Alzforum
Presentation made at the Alzforum's live webinar of December 5, 2016, titled "Is Alzheimer’s Disease a Uniquely Human Disorder?" - review additional information and recording at www.alzforum.org/
Elizabeth Head Alzforum Webinar - Dec 7, 2016Alzforum
Presentation made at the Alzforum's live webinar of December 5, 2016, titled "Is Alzheimer’s Disease a Uniquely Human Disorder?" - review additional information and recording at www.alzforum.org/
Patrick Hof Alzforum Webinar - Dec 7, 2016Alzforum
Presentation made at the Alzforum's live webinar of December 5, 2016, titled "Is Alzheimer’s Disease a Uniquely Human Disorder?" - review additional information and recording at www.alzforum.org/
Presentation made at the Alzforum's live webinar of December 5, 2016, titled "Is Alzheimer’s Disease a Uniquely Human Disorder?" - review additional information and recording at www.alzforum.org/
Presentation made at the live webinar of April 8, 2016 hosted by Alzforum - http://www.alzforum.org/webinars/webinar-pathogenic-protein-spread-lets-think-again
Dominic Walsh - A Critical Appraisal of the Pathogenic Protein Spread Hypothe...Alzforum
Presentation made April 8, 2016 at the live webinar hosted by Alzforum - http://www.alzforum.org/webinars/webinar-pathogenic-protein-spread-lets-think-again
Presentation made April 8, 2016 at the live webinar hosted by Alzforum - http://www.alzforum.org/webinars/webinar-pathogenic-protein-spread-lets-think-again
Patrik Brundin - Are Synucleinopathies Prion Diseases?Alzforum
Presentation made April 8, 2016 at the live webinar hosted by Alzforum - http://www.alzforum.org/webinars/webinar-pathogenic-protein-spread-lets-think-again
Virginia Lee - Cell-to-Cell Spread of Pathological TauAlzforum
Presentation made April 8, 2016 at the live webinar hosted by Alzforum - http://www.alzforum.org/webinars/webinar-pathogenic-protein-spread-lets-think-again
Presentation made by Dr. Cliff Brangwynne on October 30, 2015 at the Alzforum-hosted live webinar titled "Fluid Business: Could “Liquid” Protein Herald Neurodegeneration?"
More information and the recording of the session available at http://www.alzforum.org/webinars/fluid-business-could-liquid-protein-herald-neurodegeneration
Observation of Io’s Resurfacing via Plume Deposition Using Ground-based Adapt...Sérgio Sacani
Since volcanic activity was first discovered on Io from Voyager images in 1979, changes
on Io’s surface have been monitored from both spacecraft and ground-based telescopes.
Here, we present the highest spatial resolution images of Io ever obtained from a groundbased telescope. These images, acquired by the SHARK-VIS instrument on the Large
Binocular Telescope, show evidence of a major resurfacing event on Io’s trailing hemisphere. When compared to the most recent spacecraft images, the SHARK-VIS images
show that a plume deposit from a powerful eruption at Pillan Patera has covered part
of the long-lived Pele plume deposit. Although this type of resurfacing event may be common on Io, few have been detected due to the rarity of spacecraft visits and the previously low spatial resolution available from Earth-based telescopes. The SHARK-VIS instrument ushers in a new era of high resolution imaging of Io’s surface using adaptive
optics at visible wavelengths.
Earliest Galaxies in the JADES Origins Field: Luminosity Function and Cosmic ...Sérgio Sacani
We characterize the earliest galaxy population in the JADES Origins Field (JOF), the deepest
imaging field observed with JWST. We make use of the ancillary Hubble optical images (5 filters
spanning 0.4−0.9µm) and novel JWST images with 14 filters spanning 0.8−5µm, including 7 mediumband filters, and reaching total exposure times of up to 46 hours per filter. We combine all our data
at > 2.3µm to construct an ultradeep image, reaching as deep as ≈ 31.4 AB mag in the stack and
30.3-31.0 AB mag (5σ, r = 0.1” circular aperture) in individual filters. We measure photometric
redshifts and use robust selection criteria to identify a sample of eight galaxy candidates at redshifts
z = 11.5 − 15. These objects show compact half-light radii of R1/2 ∼ 50 − 200pc, stellar masses of
M⋆ ∼ 107−108M⊙, and star-formation rates of SFR ∼ 0.1−1 M⊙ yr−1
. Our search finds no candidates
at 15 < z < 20, placing upper limits at these redshifts. We develop a forward modeling approach to
infer the properties of the evolving luminosity function without binning in redshift or luminosity that
marginalizes over the photometric redshift uncertainty of our candidate galaxies and incorporates the
impact of non-detections. We find a z = 12 luminosity function in good agreement with prior results,
and that the luminosity function normalization and UV luminosity density decline by a factor of ∼ 2.5
from z = 12 to z = 14. We discuss the possible implications of our results in the context of theoretical
models for evolution of the dark matter halo mass function.
Multi-source connectivity as the driver of solar wind variability in the heli...Sérgio Sacani
The ambient solar wind that flls the heliosphere originates from multiple
sources in the solar corona and is highly structured. It is often described
as high-speed, relatively homogeneous, plasma streams from coronal
holes and slow-speed, highly variable, streams whose source regions are
under debate. A key goal of ESA/NASA’s Solar Orbiter mission is to identify
solar wind sources and understand what drives the complexity seen in the
heliosphere. By combining magnetic feld modelling and spectroscopic
techniques with high-resolution observations and measurements, we show
that the solar wind variability detected in situ by Solar Orbiter in March
2022 is driven by spatio-temporal changes in the magnetic connectivity to
multiple sources in the solar atmosphere. The magnetic feld footpoints
connected to the spacecraft moved from the boundaries of a coronal hole
to one active region (12961) and then across to another region (12957). This
is refected in the in situ measurements, which show the transition from fast
to highly Alfvénic then to slow solar wind that is disrupted by the arrival of
a coronal mass ejection. Our results describe solar wind variability at 0.5 au
but are applicable to near-Earth observatories.
Introduction:
RNA interference (RNAi) or Post-Transcriptional Gene Silencing (PTGS) is an important biological process for modulating eukaryotic gene expression.
It is highly conserved process of posttranscriptional gene silencing by which double stranded RNA (dsRNA) causes sequence-specific degradation of mRNA sequences.
dsRNA-induced gene silencing (RNAi) is reported in a wide range of eukaryotes ranging from worms, insects, mammals and plants.
This process mediates resistance to both endogenous parasitic and exogenous pathogenic nucleic acids, and regulates the expression of protein-coding genes.
What are small ncRNAs?
micro RNA (miRNA)
short interfering RNA (siRNA)
Properties of small non-coding RNA:
Involved in silencing mRNA transcripts.
Called “small” because they are usually only about 21-24 nucleotides long.
Synthesized by first cutting up longer precursor sequences (like the 61nt one that Lee discovered).
Silence an mRNA by base pairing with some sequence on the mRNA.
Discovery of siRNA?
The first small RNA:
In 1993 Rosalind Lee (Victor Ambros lab) was studying a non- coding gene in C. elegans, lin-4, that was involved in silencing of another gene, lin-14, at the appropriate time in the
development of the worm C. elegans.
Two small transcripts of lin-4 (22nt and 61nt) were found to be complementary to a sequence in the 3' UTR of lin-14.
Because lin-4 encoded no protein, she deduced that it must be these transcripts that are causing the silencing by RNA-RNA interactions.
Types of RNAi ( non coding RNA)
MiRNA
Length (23-25 nt)
Trans acting
Binds with target MRNA in mismatch
Translation inhibition
Si RNA
Length 21 nt.
Cis acting
Bind with target Mrna in perfect complementary sequence
Piwi-RNA
Length ; 25 to 36 nt.
Expressed in Germ Cells
Regulates trnasposomes activity
MECHANISM OF RNAI:
First the double-stranded RNA teams up with a protein complex named Dicer, which cuts the long RNA into short pieces.
Then another protein complex called RISC (RNA-induced silencing complex) discards one of the two RNA strands.
The RISC-docked, single-stranded RNA then pairs with the homologous mRNA and destroys it.
THE RISC COMPLEX:
RISC is large(>500kD) RNA multi- protein Binding complex which triggers MRNA degradation in response to MRNA
Unwinding of double stranded Si RNA by ATP independent Helicase
Active component of RISC is Ago proteins( ENDONUCLEASE) which cleave target MRNA.
DICER: endonuclease (RNase Family III)
Argonaute: Central Component of the RNA-Induced Silencing Complex (RISC)
One strand of the dsRNA produced by Dicer is retained in the RISC complex in association with Argonaute
ARGONAUTE PROTEIN :
1.PAZ(PIWI/Argonaute/ Zwille)- Recognition of target MRNA
2.PIWI (p-element induced wimpy Testis)- breaks Phosphodiester bond of mRNA.)RNAse H activity.
MiRNA:
The Double-stranded RNAs are naturally produced in eukaryotic cells during development, and they have a key role in regulating gene expression .
THE IMPORTANCE OF MARTIAN ATMOSPHERE SAMPLE RETURN.Sérgio Sacani
The return of a sample of near-surface atmosphere from Mars would facilitate answers to several first-order science questions surrounding the formation and evolution of the planet. One of the important aspects of terrestrial planet formation in general is the role that primary atmospheres played in influencing the chemistry and structure of the planets and their antecedents. Studies of the martian atmosphere can be used to investigate the role of a primary atmosphere in its history. Atmosphere samples would also inform our understanding of the near-surface chemistry of the planet, and ultimately the prospects for life. High-precision isotopic analyses of constituent gases are needed to address these questions, requiring that the analyses are made on returned samples rather than in situ.
Cancer cell metabolism: special Reference to Lactate PathwayAADYARAJPANDEY1
Normal Cell Metabolism:
Cellular respiration describes the series of steps that cells use to break down sugar and other chemicals to get the energy we need to function.
Energy is stored in the bonds of glucose and when glucose is broken down, much of that energy is released.
Cell utilize energy in the form of ATP.
The first step of respiration is called glycolysis. In a series of steps, glycolysis breaks glucose into two smaller molecules - a chemical called pyruvate. A small amount of ATP is formed during this process.
Most healthy cells continue the breakdown in a second process, called the Kreb's cycle. The Kreb's cycle allows cells to “burn” the pyruvates made in glycolysis to get more ATP.
The last step in the breakdown of glucose is called oxidative phosphorylation (Ox-Phos).
It takes place in specialized cell structures called mitochondria. This process produces a large amount of ATP. Importantly, cells need oxygen to complete oxidative phosphorylation.
If a cell completes only glycolysis, only 2 molecules of ATP are made per glucose. However, if the cell completes the entire respiration process (glycolysis - Kreb's - oxidative phosphorylation), about 36 molecules of ATP are created, giving it much more energy to use.
IN CANCER CELL:
Unlike healthy cells that "burn" the entire molecule of sugar to capture a large amount of energy as ATP, cancer cells are wasteful.
Cancer cells only partially break down sugar molecules. They overuse the first step of respiration, glycolysis. They frequently do not complete the second step, oxidative phosphorylation.
This results in only 2 molecules of ATP per each glucose molecule instead of the 36 or so ATPs healthy cells gain. As a result, cancer cells need to use a lot more sugar molecules to get enough energy to survive.
Unlike healthy cells that "burn" the entire molecule of sugar to capture a large amount of energy as ATP, cancer cells are wasteful.
Cancer cells only partially break down sugar molecules. They overuse the first step of respiration, glycolysis. They frequently do not complete the second step, oxidative phosphorylation.
This results in only 2 molecules of ATP per each glucose molecule instead of the 36 or so ATPs healthy cells gain. As a result, cancer cells need to use a lot more sugar molecules to get enough energy to survive.
introduction to WARBERG PHENOMENA:
WARBURG EFFECT Usually, cancer cells are highly glycolytic (glucose addiction) and take up more glucose than do normal cells from outside.
Otto Heinrich Warburg (; 8 October 1883 – 1 August 1970) In 1931 was awarded the Nobel Prize in Physiology for his "discovery of the nature and mode of action of the respiratory enzyme.
WARNBURG EFFECT : cancer cells under aerobic (well-oxygenated) conditions to metabolize glucose to lactate (aerobic glycolysis) is known as the Warburg effect. Warburg made the observation that tumor slices consume glucose and secrete lactate at a higher rate than normal tissues.
Richard's aventures in two entangled wonderlandsRichard Gill
Since the loophole-free Bell experiments of 2020 and the Nobel prizes in physics of 2022, critics of Bell's work have retreated to the fortress of super-determinism. Now, super-determinism is a derogatory word - it just means "determinism". Palmer, Hance and Hossenfelder argue that quantum mechanics and determinism are not incompatible, using a sophisticated mathematical construction based on a subtle thinning of allowed states and measurements in quantum mechanics, such that what is left appears to make Bell's argument fail, without altering the empirical predictions of quantum mechanics. I think however that it is a smoke screen, and the slogan "lost in math" comes to my mind. I will discuss some other recent disproofs of Bell's theorem using the language of causality based on causal graphs. Causal thinking is also central to law and justice. I will mention surprising connections to my work on serial killer nurse cases, in particular the Dutch case of Lucia de Berk and the current UK case of Lucy Letby.
1. J. Paul Taylor, MD, PhD
St. Jude Children’s Research Hospital
Howard Hughes Medical Institute
Dynamic RNA-protein assemblies in
neurological disease
2. Multisystem Proteinopathy – a pleiotropic
syndrome
Frontotemporal dementia
ALS
Inclusion body Myopathy
Paget’s Disease of bone
Index case
Johnston et al. Neuron 2010
Watts et al. Nature Genetics 2004
R155H mutation in VCP
Frontotemporal dementia
ALS
Inclusion body Myopathy
Paget’s Disease of bone
Index case
R155H mutation in VCP
• What is the basis of clinical pleiotropy in patients with identical VCP
mutations?
• What does this tell us about the etiologic relationship between these
distinct age-related degenerative diseases?
• What is the basis of clinical pleiotropy in patients with identical VCP
mutations?
• What does this tell us about the etiologic relationship between these
distinct age-related degenerative diseases?
3. Family 1 Family 3Family 2
Family 4 Family 5
Family 6
Family 7
Family 8
Family 9
Multisystem Proteinopathy Families
Benatar et al. Neurology 2013
4. Mutations in hnRNPA2B1 and hnRNPA1 cause MSP
hnRNP A2B1
12 aa
D290V
D302V
A2 ( 95%)
B1 ( 5%)
isoform
LCS
LCS
MS9MS9
D262V
hnRNP A1
52 aa
a (95%)
b ( 5%)
isoform
LCS
LCS
MS9MS9
D262V
Kim et al., Nature 2013
6. Gene Functional Class MSP ALS FTD IBM Paget’s
Protein
present in
pathology
VCP
Ubiquitin-dependent
segregase
✔ ✔ ✔ ✔ ✔ yes
p62/SQSTM1
Ubiquitin-dependent
autophagy
✔ ✔ ✔ ✔ ✔ yes
Optineurin
Ubiquitin-dependent
autophagy
✔ ✔ ✔ ✔ yes
Ubiquilin2
Ubiquitin-dependent
autophagy
✔ ✔ ✔ yes
TDP-43 RNA-binding protein ✔ ✔ yes
hnRNPA2B1 RNA-binding protein ✔ ✔ ✔ yes
hnRNPA1 RNA-binding protein ✔ ✔
✔
yes
hnRNPDL RNA-binding protein ✔ yes
TIA-1 RNA-binding protein ✔ ✔ ✔ yes
The Molecular Genetics
of Multisystem Proteinopathy
Taylor, Neurology 2015
7. Mutations in these same genes are found in sporadic diseas
ALS
hnRNPA1
TDP43
MATR3
FUS
p62/SQSTM1
UBQLN2
OPTN
VCP
IBM
hnRNPA2B1
MATR3
FUS
p62/SQSTM1
UBQLN2
VCP
Paget’s
hnRNPA1
hnRNPA2B1
p62/SQSTM1
UBQLN2
OPTN
VCP
FTD
TDP43
FUS
p62/SQSTM1
UBQLN2
VCP
10. hnRNPA2B1 and hnRNPA1 fibril assembly accelerated
by disease mutations
USup35
ORD M domain C domain
PrLD ORD M domain C domain
1 40 114 254 685
S V
Time (h)
G
inpellet(%)
Time (h)
H
I
J
K
M
O
Q
inpellet(%)
Time (h)
WT
D290V
Δ287-292
WT
D262V
Δ259-264
WTD290VWTD262V
0h 4h 18h
0h 4h 12h
μm
μm
inpellet(%)
Time (h)
0
20
40
60
0 1 2 3
D290Vinpellet(%)
Time (h)
0
10
20
30
0 1 2 3 4
40
50
Δ287-292inpellet(%)
Time (h)
0
10
20
30
0 1 2 3
40
hnRNPA2inpellet(%)
Time (h)
0
10
20
30
0 1 2 3
Sup35
Sup35
USup35
ORD M domain C domain
PrLD ORD M domain C domain
1 40 114 254 685
S V
Time (h)
G
inpellet(%)
Time (h)
H
I
J
K
M
O
Q
inpellet(%)
Time (h)
WT
D290V
Δ287-292
WT
D262V
Δ259-264
WTD290VWTD262V
0h 4h 18h
0h 4h 12h
μm
μm
inpellet(%)
Time (h)
0
20
40
60
0 1 2 3 4
D290Vinpellet(%)
Time (h)
0
10
20
30
0 1 2 3 4
40
50
5 6
Δ287-292inpellet(%)
Time (h)
0
10
20
30
0 1 2 3 4
40
hnRNPA2inpellet(%)
Time (h)
0
10
20
30
0 1 2 3 4
Sup35
Sup35
hnRNPA2
hnRNPA1
• Full-length wild type hnRNPA2 and hnRNPA1 fibrillize after a lag phase
• Disease mutations greatly reduce the lag phase
• Deletion of the hexapeptide “steric zipper” eliminates fibrilization Kim et al., Nature 2013
11. hnRNP incorporation into stress granules is
enhanced by mutations
hnRNPA2 D290V
hnRNPA1 D262V
Kim et al., Nature 2013
16. hnRNP protein droplets exhibit liquid-like behavior
time lapse time lapse
Molliex, Cell 2015
Oregon Green-labeled
droplets of purified hnRNPA1
GFP-tagged
stress granules
in cells
17. hnRNPs droplets are highly dynamic
Hydrogels show
no fluorescence
recovery from
sequential
bleaching even
after 16 minutes
Protein droplets
show fluorescence
recovery on a time
scale of seconds
hnRNPA1 Hydrogel
hnRNPA1 droplet
Molliex, Cell 2015
20. T [°C]
time
33
20
35
[A1-FL] (μM)
50 100 150 200 250 300
Temperature(⁰C)
10
15
20
25
30
35
150 100 75
[Ficoll] (mg/ml)
hnRNPA1 is poised at the phase boundary as we
approach physiological conditions of temperature,
concentration, salt and intracellular molecular
crowding
Mapping the phase diagram of liquid-liquid
phase separation
Individual mRNPs
m7Gppp
AAA
m7Gppp
AAA
m7Gppp
AAA
m7Gppp
AAA
m7Gppp
AAA
RNA granule
m7Gppp
AAA
m7Gppp
AAA
m7Gppp
AAA
LLPS
Hypothesis: phase separation by LCD-containing
RBPs underlies the formation of RNA granules and
related RNA-protein assemblies as well as their
liquid properties
Molliex, Cell 2015
21. Genetics and cell biology point to a
disturbance in RNA granule
dynamics in ALS and related
diseases
Enigma
Patient pathology is dominated by
deposition of RNA-binding protein
inclusions
What is the relationship between hyperassembly of persistent
stress granules and deposition of RBP aggregates?
22. hnRNPA1 WT
5 µ m
hnRNPA1 D262V
5 µ m
Merge
5 µ m
0
5
10
15
20
25
30
0 50 100 150 200 250
Temperature(⁰C)
hnRNPA1 (μM)
Diseases mutation in hnRNPA1 doesn’t
impact phase separation
hnRNPA1 WT and D262V are miscible in droplets
D262V
RRM1
RRM
2
LCS
D262V
RRM1
RRM
2
LCS
hnRNPA1
hnRNPA1-D262V
D262V
Molliex, Cell 2015