ALS Research ‘Gels’ as Studies Tie Disparate Genetic Factors Together

Run-on repeats of a hexanucleotide sequence in the C9ORF72 gene are the most common cause of familial amyotrophic lateral sclerosis and frontotemporal dementia, but do they relate to other ALS/FTD mutations? According to two studies published in Cell on October 20, arginine-containing repeat dipeptides translated from the C9ORF72 expansions buddy up with proteins that have low-complexity domains and tend to form liquid organelles. These include the ALS/FTD culprits TDP-43 and FUS. The dipeptide repeats transformed these liquid droplets—including stress granules and the nucleolus—into molasses, preventing the free movement of proteins in and out of the organelles. This, in turn, disrupts fundamental cellular processes, including translation and protein transport, claim the authors.

“These papers are really exciting,” commented Gene Yeo of the University of California, San Diego. “There is now a convergence in the downstream molecular mechanisms that cause ALS/FTD from mutations in RNA-binding proteins and C9ORF72.” 

Droplet Instigators.

The nucleolar protein NPM1 (left) formed liquid droplets when mixed with polyGR (middle) or polyPR (right), two types of dipeptide repeat derived from C9ORF72 hexanucleotide repeat expansions. [Image courtesy of Lee et al., Cell 2016.]

One study, led by J. Paul Taylor at St. Jude Children’s Research Hospital in Memphis, Tennessee, uncovered how C9 dipeptide repeat proteins (DPRs) affected the dynamics and function of multiple membrane-less organelles. The other, led by Steven McKnight at University of Texas Southwestern Medical Center in Dallas, focused on the biophysical interactions between the dipeptides and their targets. Together, the studies raise the possibility of a broad pathological mechanism shared between carriers of the C9ORF72 expansion and other disease-causing mutations. 

While a healthy person harbors between two and 23 repeats of the hexanucleotide sequence GGGGCC in their C9ORF72 gene, people with ALS or FTD can have hundreds to thousands. These sequences are then translated in both the sense and antisense directions to yield five different dipeptide repeats (DPRs): glycine-alanine (GA), glycine-arginine (GR), proline-arginine (PR), proline-alanine (PA), or glycine-proline (GP). Of these, the two arginine-containing DPRs—GR and PR—have the biggest wrap sheets. They reportedly clog the nucleolus and bungle RNA biogenesis, form toxic nuclear aggregates and stress granules in neurons, block traffic between the nucleus and cytoplasm, and kill flies, just to name a few (see Aug 2014 news; Dec 2014 news; Aug 2015 news). 

Both Taylor and McKnight wanted to understand how these dipeptides might harm neurons, so both carried out screens to find proteins with which the peptides interacted. In Taylor’s lab, co-first authors Kyung-Ha Lee and Peipei Zhang and colleagues expressed 47-50 copies of each repeat sequence in HEK293T cells, immunoprecipitated the dipeptides, and analyzed any proteins that came along for the ride via liquid chromatography and mass spectrometry. While GA, GP, and PA repeats largely came up solo, the two arginine-containing dipeptides had 196 different partners, 81 of which associated with both dipeptides. Nearly 70 percent of these GR/PR associates—which included the ALS-linked proteins TDP-43, FUS, hnRNPA1, and hnRNPA2B1—contained low-complexity domains (LCDs).

Taylor and others had previously reported that these domains facilitated the formation of membrane-less organelles and might be hotbeds for protein aggregation (see Oct 2015 news; Oct 2015 webinar). Knocking down expression of most of these interactors in flies either rescued or worsened the toxic effects of the GR dipeptide repeats in the insects, indicating that the interactions played a meaningful role in facilitating DPR toxicity.

The researchers next focused in on how GR/PR dipeptide repeats affected different membrane-less organelles. A common theme emerged: Because the arginine-rich repeats interacted strongly with LCD-containing proteins in each organelle, they essentially “gelled” the organelles. For example, in the nucleolus—a structure within the nucleus where ribosomal RNA is generated, processed, and released—the polyGR/PR dipeptides associated with a protein called NPM1. By associating with other, physiological arginine-rich proteins, NPM1 normally orchestrates the formation of the liquid-like “granular component” of the nucleolus, where rRNA biogenesis takes place. When C9ORF72 GR/PRs were added to the mix, they outcompeted NPM1’s other partners, kicking droplet formation into overdrive and rendering the granular component more viscous. This essentially halted the movement of proteins in and out of the structure (see image above).

This, in turn, deprived the cell of rRNA, which remained trapped in the granular component of the nucleolus. The researchers reported a similar gelling phenomenon in stress granules. They also spotted the GR/PR peptides in other membrane-less nuclear organelles, including nuclear speckles, which are enriched with splicing and transcription factors, and in Cajal bodies, which host the spliceosome. Key proteins in each of these structures had popped up in the original GR/PR binding screen. The resulting loss of protein mobility profoundly affected the functions of these organelles.

“When GR/PR peptides infiltrate these organelles, the liquid goes from the viscosity of honey on the table to honey in the refrigerator,” Taylor explained. He pointed out that roughly a third of proteins contain LCDs, many of which may facilitate liquid organelle formation by associating with arginine-rich sequences. He speculated that such liquid phase separation likely underlies a vast array of dynamic biological processes, such as rapid clustering of receptors on membranes. “Nature would be hard-pressed to design a more potent cellular poison than arginine-rich polymers,” Taylor said.

For his part, McKnight in Dallas also screened for partners of dipeptide repeats, and pulled out many of the same hits as did Taylor’s group, including a preponderance of LCD-containing proteins. In addition, the Dallas group found intermediate filaments that bound polyPR. These filaments, which include neurofilaments and vimentin, help form the cellular cytoskeleton necessary for transport of proteins throughout the cell. 

Rather than looking at specific membrane-less organelles that might be affected by these interactions, co-first authors Yi Lin and Eiichiro Mori investigated the fundamental mechanisms by which PR peptides changed the behavior of LCD-containing proteins. Previous studies from McKnight’s lab had indicated that under physiological conditions, the LCDs of numerous RNA-binding proteins, including FUS, polymerized into fibers of cross β-sheets, and that this facilitates the formation of liquid organelles (see Kato et al., 2012; Han et al., 2012). Therefore, the researchers started by asking whether LCD-containing proteins needed to polymerize to bind PR dipeptides. The researchers treated liquid protein droplets or organelles with the aliphatic alcohol 1,6-hexanediol, which depolymerizes the proteins and dissolves these structures. They found that this depolymerization abolished the association between polyPR peptides and LCD containing proteins, including FUS. 

Deadly Dew? Vimentin (left) bound droplets of polyPR peptides (middle) or droplets of FUS (right). Researchers proposed the PR peptides could usurp vimentin’s contacts with RNA granules. [Image courtesy of Lin et al., Cell 2016.]

Interestingly, among three RNA-binding proteins—hnRNPA1, hnRNPA2, and hnRNPDL—mutations that lead to ALS, FTD, and muscular dystrophy occur in the same aspartic acid within each protein’s LCD. The researchers found that these mutants formed more stable polymers than did the normal versions, and that in the case of the D290V mutant of hnRNPA2, the mutant bound to PR peptide repeats more strongly. Conversely, an hnRNPA2 mutant that could not form polymers failed to bind polyPR. Together, these findings indicated that PR peptides only associated with polymerized LCD-containing proteins, which would explain their uncanny ability to disrupt liquid organelles.

Finally, Lin and colleagues examined the association between polyPR peptides and intermediate filaments. These proteins contain LCDs in the amino terminal (“head”) domain, as well as their C-terminal (“tail”) domain, which flank a central α-helical portion. These LCDs polymerized with each other to form filaments, and, much like liquid organelles, were disrupted by aliphatic alcohols. The researchers reported that PR peptide repeats bound to these LCDs along polymerized filaments, forming a series of regularly spaced knobs (see image above). They speculated that these PR peptide globs could perturb the critical functions of these filaments. For example, neurofilaments near the synapse associate with RNA granules. If PR aggregates outcompeted RNA granules for binding to neurofilament, it would prevent localized protein translation near the synapse, they proposed.

Yeo believes the findings support the idea that stress granule dynamics are disrupted in these diseases. The studies also expand the list of potentially affected structures, such as the nucleolus and Cajal bodies, he said. Yeo recently reported that the D290V mutant of hnRNPA2B1 affected stress granules, but that it also triggered cell death under low stress conditions, in which those granules did not form. The mutation also fouled up alternative splicing, he reported (see Martinez et al., 2016). The new findings introduce the possibility that disruption of other membrane-less organelles, such as the spliceosome, could explain D290V toxicity, Yeo said.

“These papers raise critical questions regarding the mechanism of DPR toxicity in ALS,” commented Nicolas Fawzi of Brown University in Providence, Rhode Island. “For example, the structural details of how DPRs interact with LCDs of other disease-related RNA-binding proteins are now of prime importance,” he wrote. “It will also be interesting to examine the interaction of DPRs with TDP-43, which can self-assemble via α-helical structure present with its LCD.” Fawzi’s previous work described the formation of liquid droplets by both FUS and TDP-43 proteins (see Oct 2015 news on Burke et al., 2015; Sep 2016 news on Conicella et al., 2016). Fawzi also wondered whether disruption of LCDs could underlie cases of sporadic ALS as well.

Do these findings move researchers closer to developing treatments for ALS or FTD? Taylor thinks so. “Liquid phase separations that form membrane-less organelles are highly regulated, tunable processes,” he told Alzforum. “For every factor that promotes assembly, another promotes disassembly, so there may be antagonistic relationships we can exploit.”—Jessica Shugart

Comments

  1. These papers convincingly argue that toxicity due to C9ORF72 GGGGCC repeat expansions arises from arginine dipeptide repeats (DPRs) interactions with proteins containing low-complexity domain (LCDs) inside ribonucleoprotein granule membrane-less organelles. Intriguingly, many of these LCD proteins themselves form inclusions in, and contain mutations causing, ALS (TDP-43, FUS, hnRNP A2). Using pull-down proteomic approaches, in-cell granule localization, and in vitro reconstitution of liquid droplet and hydrogel models of RNP granules, the authors demonstrate that arginine-rich DPRs interact with low-complexity domains of RNA-binding proteins and other granule components, and even intermediate filament disordered domains. These interactions are shown to alter dynamics of these subcellular membrane-less organelles. Though GGGGCC repeat expansions code for non-arginine peptides as well, Lee et al. demonstrate that arginine is required for granule interactions and toxicity. These papers lead to several critical unanswered mechanistic questions regarding DPRs and ALS.

    1) What is the role of arginine-rich regions and RG/RGG repeats natively present in ALS-associated LCD proteins such as FUS? Our previous work demonstrates that although the low-complexity domain of FUS (residues 1-163), which lacks arginine and other positively charged residues, is alone competent for liquid-liquid phase separation into models of RNP granules, full-length FUS, which includes three RGG repeat regions, phase separates at much lower sub-physiological concentration ranges (Burke et al.,  2015; Oct 2015 news). Additionally, several reports demonstrate that these regions are required for toxicity. Therefore, interactions between RGG regions and low-complexity domains natively present may be outcompeted by arginine-rich polymers and altered by ALS-associated mutations. And yet, the atomic structural details of DPR-LCD interaction are still unknown. It will be critical to conclusively determine where physiological RNP LCDs lie on the continuum between intrinsic disorder and ordered cross-β structure, and how DPRs alter this structure. It will also be a top priority to examine the interaction of DPRs with TDP-43, which can self-assemble via α-helical structure present within its LCD (Conicella et al., 2016; Sep 2016 news). 

    2) What is the mechanism of the observed toxicity and the suppressor/enhancer phenotypes observed? Given that several LCD-containing proteins that interact with arginine DPRs are included in the list of modifiers, it will be exciting to determine how altering LCD protein levels modulates toxicity.

    3) Finally, does disruption of LCD proteins in RNPs explain not only C9ORF72 ALS, as suggested by these papers, but rather all ALS, including those with no known genetic component? The answer to this question, which may be found using the approaches demonstrated in these papers, will be important for designing strategies to prevent ALS or halt its progression. 

    References:

    Burke KA, Janke AM, Rhine CL, Fawzi NL. Residue-by-Residue View of In Vitro FUS Granules that Bind the C-Terminal Domain of RNA Polymerase II. Mol Cell. 2015 Oct 15;60(2):231-41. Epub 2015 Oct 8 PubMed.

    Conicella AE, Zerze GH, Mittal J, Fawzi NL. ALS Mutations Disrupt Phase Separation Mediated by α-Helical Structure in the TDP-43 Low-Complexity C-Terminal Domain. Structure. 2016 Sep 6;24(9):1537-49. Epub 2016 Aug 18 PubMed.

    View all comments by Nicolas Fawzi

  2. These two papers present a comprehensive addition to our knowledge of the mechanisms of action of C9ORF72 dipeptide repeats. Studying other protein aggregates, investigators identified a plethora of protein interactions, which lead to the general consensus that the key to preventing toxicity caused by protein aggregates is to prevent their formation or to reduce their abundance. The original set of manuscripts on the C9ORF72 dipeptide repeats suggested that interactions with nucleoli or the nuclear pores were a main target of action. The current manuscripts confirm these interactions, but convey the much more compelling case that the dipeptide repeats interact with many, many proteins within the cell, potentially causing injury through a multiplicity of mechanisms.

    View all comments by Benjamin Wolozin

  3. Since the discovery of C9ORF72 hexanucleotide expansions in patients with ALS and frontotemporal dementia (Dejesus-Hernandez et al., 2011; Renton et al., 2011), controversy has ensued about the cellular mechanisms that drive neurodegeneration in this patient cohort (Gitler and Tsuiji, 2016). While some data suggest that haploinsufficiency could contribute to disease (O'Rourke et al., 2016; Sivadasan et al., 2016), the vast majority of studies instead support a dominant gain of function as the major causal player in disease pathogenesis. Cell biologists have been roughly divided into two camps: one that suggests that transcribed C9ORF72 RNA begets dysfunction (Zhang et al., 2015), whereas the other argues that unconventionally translated dipeptides give rise to disease (Zu et al., 2011). Initial mammalian cell culture studies from Steven McKnight’s lab convincingly demonstrated that exogenously applied arginine-rich dipeptides are sufficient to kill cells, likely via dysfunction of the nucleolus (Kwon et al., 2014). Screens from Aaron Gitler and Paul Taylor’s labs further suggested that nuclear import/export could be impaired by arginine-rich dipeptides, but not by other dipeptides produced from the C9ORF72 locus (Freibaum et al., 2015; Jovičić et al., 2015), although the Taylor study did not rule out a role for expanded RNA transcripts in driving toxicity.

    This new Cell paper from Paul Taylor and colleagues is an elegant piece of work that combines proteomics, Drosophila screening, in vitro biochemistry, and live cell dynamics to show that arginine-rich dipeptides “poison” a vast array of membraneless organelles that arise by liquid-liquid phase separation from the nucleoplasm and cytoplasm (Brangwynne et al., 2009; Weber and Brangwynne, 2012). This seminal paper can be appreciated from two different angles:

    1. From a therapeutic perspective, because it suggests that drug development should aim to reduce arginine-rich dipeptide production in patients with C9ORF72 hexanucleotide repeat expansions.
    2. From a basic cell biology standpoint, because it gives further credence to the hypothesis that electrostatics and multivalent blocked-charge motifs are of critical importance to the assembly and regulation of diverse membraneless organelles, e.g., nucleoli, splicing speckles, Cajal bodies, stress granules, paraspeckles (Brangwynne et al., 2015; Nott et al., 2015; Pak et al., 2016). 

    While I applaud the importance of the findings from a cell biology perspective (I know the term “tour de force” is used too often by scientific commentators, but nevertheless, I think the phrase is applicable in this case), I wish to add some caveats regarding the relevance to disease pathogenesis, which I believe are lacking in the Alzforum perspective. First, I must emphasize that the authors do not argue that arginine-rich dipeptides are the only contributing factor to C9ORF-ALS/FTD pathogenesis.

    It is now quite clear from numerous studies that large amounts of positively charged dipeptides are unequivocally “bad” for cells. This is likely due to the fact that arginine-rich dipeptides compete with endogenous RGG motif-containing proteins for electrostatic interactions with negatively charged and phosphorylated proteins enriched in membraneless organelles. However, it is not clear to me how accurately current cell and organismal models recapitulate critical features of C9ORF-ALS/FTD. Exhaustive immunohistochemical studies have shown neuronal loss correlates with TDP-43 pathology but not dipeptide pathology (Mackenzie et al., 2015). Moreover, unlike all dipeptide models, most human dipeptide pathology is observed in the cytoplasm and not the nucleus, (Mackenzie et al., 2015). Further, poly-GA and poly-GP inclusions are far more abundant than poly-PR or poly-GR accumulations (Mackenzie et al., 2015). Finally, when dipeptides are observed in the nucleus, they appear to localize to a perinucleolar compartment and not the granular component of the nucleolus (Schludi et al., 2015; Vatsavayai et al., 2016). With these pathological observations in mind, I think it is important that we continue to consider all possible mechanisms (haploinsufficiency, RNA toxicity, arginine dipeptide toxicity, GA dipeptide toxicity) in the pathogenesis of C9ORF-ALS/FTD. Going forward, some important topics/questions come to mind that will be essential to explore for this rapidly evolving research field (i.e., the relationship between physiological phase separation and pathological protein aggregation and neurodegeneration):

    1. To what degree does each proposed pathological mechanism give rise to TDP-43 pathology? Do dipeptides, C9ORF72 haploinsufficiency, or RNA toxicity directly drive the accumulation of TDP-43 in the cytoplasm of neurons? Are these upstream factors the “trigger” and TDP-43 the “bullet” (i.e., TDP-43 is initially induced by C9ORF72-related pathology to form self-propagating prions that then spread throughout the brain to drive progressive neurodegeneration (Sanders et al., 2016)), or is TDP-43 accumulation merely a correlate irrelevant to disease pathogenesis? It is interesting that a recent paper from Bill Seeley’s lab shows that neurodegeneration can occur in the absence of overt TDP-43 pathology, at least in certain patients (Vatsavayai et al., 2016). 
    2. Why is TDP-43 so prone to forming amyloid-like cytoplasmic aggregates, relative to other RNA-binding proteins with prion-like and low-complexity domains? For example, G3BP1 is thought to be a “condensate” that allows mRNPs to self-associate into membraneless organelles called stress granules (Kedersha et al., 2013; 2016), but does not accumulate in large fibrils in ALS/FTD.
    3. If arginine-rich dipeptides are killing neurons, how do they do so in patients? Pathology studies do not show massive accumulations of these dipeptides in nucleoli and other nuclear bodies. Is it possible that this is simply a detection problem (i.e., antibodies are not sensitive enough or epitopes are buried)? Perhaps prolonged, low-level expression of dipeptides slowly alters membraneless organelle dynamics until “catastrophes” occur, whereby proteins like TDP-43 undergo a liquid-to-solid phase transition and become a self-propagating amyloid (or prion) (Sanders et al., 2016; Weber and Brangwynne, 2012). 
    4. If proteins in membraneless organelles are, in fact, in a precarious biophysical state and are prone to undergoing liquid-to-solid phase transitions (Weber and Brangwynne, 2012), why do we so rarely observe nuclear accumulations of solid fibril-like structures in patient brains relative to cytoplasmic aggregates? What is allowing the fluidity of these structures to be maintained over years and decades?
    5. With respect to the current paper from Taylor’s group, why does knockdown of ostensibly similar proteins lead to divergent outcomes (enhancers versus suppressors)? On cursory examination, there does not appear to be any logic to this at the level of amino acid composition (e.g., isoelectric point). This was the most fascinating aspect of the paper to me, as it suggests that there is still a wealth of information to be discovered from the study of proteins that form these organelles. It is not as simple as “electrostatics.” Moreover, I wish to emphasize the incredible percentage of modifiers from the biased screen (85 percent!). Nevertheless, it would have been nice to see a larger control panel of genes (e.g., generic proteins or even better yet, RNA-binding proteins without LCS domains) to get a good sense of the specificity of enhancer/suppressor phenotypes to the proteomic hits. Two controls were used, V60100 and w118, which showed ~25 percent and ~48 percent viability, respectively. Thus, it appears that the genetic system used in the paper may be especially prone to perturbation. A weakness of the study was that the cutoff to define an enhancer or suppressor was somewhat arbitrary based on the number of controls. 
    6. What accounts for the differences between cell culture/organismal models and patients with regard to sub-nuclear accumulations of arginine-rich dipeptides? Is it length or concentration dependent? This is a fascinating and vexing problem!
    7. If it is true that C9ORF72 dipeptides disrupt nuclear bodies and lead to neuronal dysfunction, why is it that pathology spreads in a hierarchical and stereotyped manner in C9ORF-ALS/FTD patients? The most parsimonious explanation for this observation is the release and uptake of a toxic factor (e.g., a prion) in a neuronal network (Sanders et al., 2016). Do dipeptides act like prions? The vast majority of studies do not as yet support this hypothesis, although one recent study demonstrated transcellular spread of dipeptides, but not “seeding” in the recipient cells (Westergard et al., 2016). 
    8. Finally, I think many questions are raised about the dynamic interplay between these numerous RNP bodies in the cells. Future work must determine what regulates shuttling of individual proteins from one compartment to the next, and how individual proteins are targeted to specific organelles despite similar amino acid compositions (Nott et al., 2015). 

    Like any great paper, these two lead to more questions than they answer. Needless to say, it is an exciting time for the field fof phase separation and neurodegeneration!

    References:

    Brangwynne CP, Eckmann CR, Courson DS, Rybarska A, Hoege C, Gharakhani J, Jülicher F, Hyman AA. Germline P granules are liquid droplets that localize by controlled dissolution/condensation. Science. 2009 Jun 26;324(5935):1729-32. Epub 2009 May 21 PubMed.

    Brangwynne CP, Tompa P, Pappu RV. Polymer physics of intracellular phase transitions. Nature Physics 11, 899–904. (2015)

    DeJesus-Hernandez M, Mackenzie IR, Boeve BF, Boxer AL, Baker M, Rutherford NJ, Nicholson AM, Finch NA, Flynn H, Adamson J, Kouri N, Wojtas A, Sengdy P, Hsiung GY, Karydas A, Seeley WW, Josephs KA, Coppola G, Geschwind DH, Wszolek ZK, Feldman H, Knopman DS, Petersen RC, Miller BL, Dickson DW, Boylan KB, Graff-Radford NR, Rademakers R. Expanded GGGGCC hexanucleotide repeat in noncoding region of C9ORF72 causes chromosome 9p-linked FTD and ALS. Neuron. 2011 Oct 20;72(2):245-56. Epub 2011 Sep 21 PubMed.

    Freibaum BD, Lu Y, Lopez-Gonzalez R, Kim NC, Almeida S, Lee KH, Badders N, Valentine M, Miller BL, Wong PC, Petrucelli L, Kim HJ, Gao FB, Taylor JP. GGGGCC repeat expansion in C9orf72 compromises nucleocytoplasmic transport. Nature. 2015 Sep 3;525(7567):129-33. Epub 2015 Aug 26 PubMed.

    Gitler AD, Tsuiji H. There has been an awakening: Emerging mechanisms of C9orf72 mutations in FTD/ALS. Brain Res. 2016 Sep 15;1647:19-29. Epub 2016 Apr 6 PubMed.

    Jovičić A, Mertens J, Boeynaems S, Bogaert E, Chai N, Yamada SB, Paul JW 3rd, Sun S, Herdy JR, Bieri G, Kramer NJ, Gage FH, Van Den Bosch L, Robberecht W, Gitler AD. Modifiers of C9orf72 dipeptide repeat toxicity connect nucleocytoplasmic transport defects to FTD/ALS. Nat Neurosci. 2015 Sep;18(9):1226-9. PubMed.

    Kedersha N, Ivanov P, Anderson P. Stress granules and cell signaling: more than just a passing phase?. Trends Biochem Sci. 2013 Oct;38(10):494-506. Epub 2013 Sep 10 PubMed.

    Kedersha N, Panas MD, Achorn CA, Lyons S, Tisdale S, Hickman T, Thomas M, Lieberman J, McInerney GM, Ivanov P, Anderson P. G3BP-Caprin1-USP10 complexes mediate stress granule condensation and associate with 40S subunits. J Cell Biol. 2016 Mar 28;212(7):845-60. PubMed.

    Kwon I, Xiang S, Kato M, Wu L, Theodoropoulos P, Wang T, Kim J, Yun J, Xie Y, McKnight SL. Poly-dipeptides encoded by the C9orf72 repeats bind nucleoli, impede RNA biogenesis, and kill cells. Science. 2014 Sep 5;345(6201):1139-45. Epub 2014 Jul 31 PubMed.

    Lee KH, Zhang P, Kim HJ, Mitrea DM, Sarkar M, Freibaum BD, Cika J, Coughlin M, Messing J, Molliex A, Maxwell BA, Kim NC, Temirov J, Moore J, Kolaitis RM, Shaw TI, Bai B, Peng J, Kriwacki RW, Taylor JP. C9orf72 Dipeptide Repeats Impair the Assembly, Dynamics, and Function of Membrane-Less Organelles. Cell. 2016 Oct 20;167(3):774-788.e17. PubMed.

    Mackenzie IR, Frick P, Grässer FA, Gendron TF, Petrucelli L, Cashman NR, Edbauer D, Kremmer E, Prudlo J, Troost D, Neumann M. Quantitative analysis and clinico-pathological correlations of different dipeptide repeat protein pathologies in C9ORF72 mutation carriers. Acta Neuropathol. 2015 Dec;130(6):845-61. Epub 2015 Sep 15 PubMed.

    Nott TJ, Petsalaki E, Farber P, Jervis D, Fussner E, Plochowietz A, Craggs TD, Bazett-Jones DP, Pawson T, Forman-Kay JD, Baldwin AJ. Phase transition of a disordered nuage protein generates environmentally responsive membraneless organelles. Mol Cell. 2015 Mar 5;57(5):936-47. PubMed.

    O'Rourke JG, Bogdanik L, Yáñez A, Lall D, Wolf AJ, Muhammad AK, Ho R, Carmona S, Vit JP, Zarrow J, Kim KJ, Bell S, Harms MB, Miller TM, Dangler CA, Underhill DM, Goodridge HS, Lutz CM, Baloh RH. C9orf72 is required for proper macrophage and microglial function in mice. Science. 2016 Mar 18;351(6279):1324-9. PubMed.

    Pak CW, Kosno M, Holehouse AS, Padrick SB, Mittal A, Ali R, Yunus AA, Liu DR, Pappu RV, Rosen MK. Sequence Determinants of Intracellular Phase Separation by Complex Coacervation of a Disordered Protein. Mol Cell. 2016 Jul 7;63(1):72-85. PubMed.

    Renton AE, Majounie E, Waite A, Simón-Sánchez J, Rollinson S, Gibbs JR, Schymick JC, Laaksovirta H, van Swieten JC, Myllykangas L, Kalimo H, Paetau A, Abramzon Y, Remes AM, Kaganovich A, Scholz SW, Duckworth J, Ding J, Harmer DW, Hernandez DG, Johnson JO, Mok K, Ryten M, Trabzuni D, Guerreiro RJ, Orrell RW, Neal J, Murray A, Pearson J, Jansen IE, Sondervan D, Seelaar H, Blake D, Young K, Halliwell N, Callister JB, Toulson G, Richardson A, Gerhard A, Snowden J, Mann D, Neary D, Nalls MA, Peuralinna T, Jansson L, Isoviita VM, Kaivorinne AL, Hölttä-Vuori M, Ikonen E, Sulkava R, Benatar M, Wuu J, Chiò A, Restagno G, Borghero G, Sabatelli M, ITALSGEN Consortium, Heckerman D, Rogaeva E, Zinman L, Rothstein JD, Sendtner M, Drepper C, Eichler EE, Alkan C, Abdullaev Z, Pack SD, Dutra A, Pak E, Hardy J, Singleton A, Williams NM, Heutink P, Pickering-Brown S, Morris HR, Tienari PJ, Traynor BJ. A hexanucleotide repeat expansion in C9ORF72 is the cause of chromosome 9p21-linked ALS-FTD. Neuron. 2011 Oct 20;72(2):257-68. Epub 2011 Sep 21 PubMed.

    Sanders DW, Kaufman SK, Holmes BB, Diamond MI. Prions and Protein Assemblies that Convey Biological Information in Health and Disease. Neuron. 2016 Feb 3;89(3):433-48. PubMed.

    Schludi MH, May S, Grässer FA, Rentzsch K, Kremmer E, Küpper C, Klopstock T, German Consortium for Frontotemporal Lobar Degeneration, Bavarian Brain Banking Alliance, Arzberger T, Edbauer D. Distribution of dipeptide repeat proteins in cellular models and C9orf72 mutation cases suggests link to transcriptional silencing. Acta Neuropathol. 2015 Oct;130(4):537-55. Epub 2015 Jun 18 PubMed.

    Sivadasan R, Hornburg D, Drepper C, Frank N, Jablonka S, Hansel A, Lojewski X, Sterneckert J, Hermann A, Shaw PJ, Ince PG, Mann M, Meissner F, Sendtner M. C9ORF72 interaction with cofilin modulates actin dynamics in motor neurons. Nat Neurosci. 2016 Dec;19(12):1610-1618. Epub 2016 Oct 10 PubMed.

    Vatsavayai SC, Yoon SJ, Gardner RC, Gendron TF, Vargas JN, Trujillo A, Pribadi M, Phillips JJ, Gaus SE, Hixson JD, Garcia PA, Rabinovici GD, Coppola G, Geschwind DH, Petrucelli L, Miller BL, Seeley WW. Timing and significance of pathological features in C9orf72 expansion-associated frontotemporal dementia. Brain. 2016 Dec;139(Pt 12):3202-3216. Epub 2016 Oct 22 PubMed.

    Weber SC, Brangwynne CP. Getting RNA and protein in phase. Cell. 2012 Jun 8;149(6):1188-91. PubMed.

    Westergard T, Jensen BK, Wen X, Cai J, Kropf E, Iacovitti L, Pasinelli P, Trotti D. Cell-to-Cell Transmission of Dipeptide Repeat Proteins Linked to C9orf72-ALS/FTD. Cell Rep. 2016 Oct 11;17(3):645-652. PubMed.

    Zhang K, Donnelly CJ, Haeusler AR, Grima JC, Machamer JB, Steinwald P, Daley EL, Miller SJ, Cunningham KM, Vidensky S, Gupta S, Thomas MA, Hong I, Chiu SL, Huganir RL, Ostrow LW, Matunis MJ, Wang J, Sattler R, Lloyd TE, Rothstein JD. The C9orf72 repeat expansion disrupts nucleocytoplasmic transport. Nature. 2015 Sep 3;525(7567):56-61. Epub 2015 Aug 26 PubMed.

    Zu T, Gibbens B, Doty NS, Gomes-Pereira M, Huguet A, Stone MD, Margolis J, Peterson M, Markowski TW, Ingram MA, Nan Z, Forster C, Low WC, Schoser B, Somia NV, Clark HB, Schmechel S, Bitterman PB, Gourdon G, Swanson MS, Moseley M, Ranum LP. Non-ATG-initiated translation directed by microsatellite expansions. Proc Natl Acad Sci U S A. 2011 Jan 4;108(1):260-5. Epub 2010 Dec 20 PubMed.

    View all comments by David Sanders

Make a Comment

To make a comment you must login or register.

References

News Citations

  1. C9ORF72 Killer Dipeptides Clog the Nucleolus
  2. Live-Cell Studies Blame Arginine Peptides for C9ORF72’s Crimes
  3. ALS Gene Repeats Obstruct Traffic Between Nucleus and Cytoplasm
  4. Do Membraneless Organelles Host Fibril Nucleation?
  5. Helical Tail Holds Sway Over TDP-43 Packaging

Webinar Citations

  1. Fluid Business: Could “Liquid” Protein Herald Neurodegeneration?

Paper Citations

  1. Kato M, Han TW, Xie S, Shi K, Du X, Wu LC, Mirzaei H, Goldsmith EJ, Longgood J, Pei J, Grishin NV, Frantz DE, Schneider JW, Chen S, Li L, Sawaya MR, Eisenberg D, Tycko R, McKnight SL. Cell-free formation of RNA granules: low complexity sequence domains form dynamic fibers within hydrogels. Cell. 2012 May 11;149(4):753-67. PubMed.
  2. Han TW, Kato M, Xie S, Wu LC, Mirzaei H, Pei J, Chen M, Xie Y, Allen J, Xiao G, McKnight SL. Cell-free formation of RNA granules: bound RNAs identify features and components of cellular assemblies. Cell. 2012 May 11;149(4):768-79. PubMed.
  3. Martinez FJ, Pratt GA, Van Nostrand EL, Batra R, Huelga SC, Kapeli K, Freese P, Chun SJ, Ling K, Gelboin-Burkhart C, Fijany L, Wang HC, Nussbacher JK, Broski SM, Kim HJ, Lardelli R, Sundararaman B, Donohue JP, Javaherian A, Lykke-Andersen J, Finkbeiner S, Bennett CF, Ares M Jr, Burge CB, Taylor JP, Rigo F, Yeo GW. Protein-RNA Networks Regulated by Normal and ALS-Associated Mutant HNRNPA2B1 in the Nervous System. Neuron. 2016 Nov 23;92(4):780-795. Epub 2016 Oct 20 PubMed.
  4. Burke KA, Janke AM, Rhine CL, Fawzi NL. Residue-by-Residue View of In Vitro FUS Granules that Bind the C-Terminal Domain of RNA Polymerase II. Mol Cell. 2015 Oct 15;60(2):231-41. Epub 2015 Oct 8 PubMed.
  5. Conicella AE, Zerze GH, Mittal J, Fawzi NL. ALS Mutations Disrupt Phase Separation Mediated by α-Helical Structure in the TDP-43 Low-Complexity C-Terminal Domain. Structure. 2016 Sep 6;24(9):1537-49. Epub 2016 Aug 18 PubMed.

Further Reading

Papers

  1. Chong PA, Forman-Kay JD. A New Phase in ALS Research. Structure. 2016 Sep 6;24(9):1435-6. PubMed.

Primary Papers

  1. Lee KH, Zhang P, Kim HJ, Mitrea DM, Sarkar M, Freibaum BD, Cika J, Coughlin M, Messing J, Molliex A, Maxwell BA, Kim NC, Temirov J, Moore J, Kolaitis RM, Shaw TI, Bai B, Peng J, Kriwacki RW, Taylor JP. C9orf72 Dipeptide Repeats Impair the Assembly, Dynamics, and Function of Membrane-Less Organelles. Cell. 2016 Oct 20;167(3):774-788.e17. PubMed.
  2. Lin Y, Mori E, Kato M, Xiang S, Wu L, Kwon I, McKnight SL. Toxic PR Poly-Dipeptides Encoded by the C9orf72 Repeat Expansion Target LC Domain Polymers. Cell. 2016 Oct 20;167(3):789-802.e12. PubMed.

Annotate

To make an annotation you must Login or Register.