Mutation of the ALS/FTD-associated RNA-binding protein FUS affects axonal development.

Aberrant condensation and localisation of the RNA-binding protein (RBP) fused in sarcoma (FUS) occur in variants of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Changes in RBP function are commonly associated with changes in axonal cytoskeletal organisation and branching in neurodevelopmental disorders. Here, we asked whether branching defects also occur in vivo in a model of FUS-associated disease. We use two reported Xenopus models of ALS/FTD (of either sex), the ALS-associated mutant FUS(P525L) and a mimic of hypomethylated FUS, FUS(16R). Both mutants strongly reduced axonal complexity in vivo. We also observed an axon looping defect for FUS(P525L) in the target area, which presumably arises due to errors in stop cue signalling. To assess whether loss of axon complexity also had a cue-independent component, we assessed axonal cytoskeletal integrity in vitro Using a novel combination of fluorescence and atomic force microscopy, we found that mutant FUS reduced actin density in the growth cone, altering its mechanical properties. Therefore, FUS mutants may induce defects during early axonal development.Significance statement This study demonstrates that mutation of the ALS/FTD (amyotrophic lateral sclerosis/frontotemporal dementia)-associated RNA-binding protein Fused in Sarcoma (FUS) can result in changes in axonal development. These changes occur both axon-autonomously in cytoskeletal organisation during axon extension and context-dependently during axonal branching. This indicates pre-symptomatic, developmental changes in axonal organisation may occur in familial disease variants.

Comparative Evaluation of Microbial Flora of Endodontic Origin in Teeth with Endo-Perio Lesions.

Background: Endodontic microbial flora plays a pivotal role in the development and persistence of periodontal endodontic lesions (PELs). Understanding the composition and prevalence of microbial species in PELs is essential for effective treatment strategies. Materials and Methods: Microbial samples were collected from 50 teeth diagnosed with PELs. Sterile paper points were used to obtain samples from the root canals. Deoxyribonucleic acid (DNA) was extracted and subjected to polymerase chain reaction (PCR) amplification of the 16S ribosomal RNA (rRNA) gene to identify bacterial species. The obtained data were analyzed using statistical methods. Results: The microbial analysis revealed a diverse range of bacterial species in PELs. The most prevalent species were Porphyromonas gingivalis (32.5%), Treponema denticola (28.0%), and Fusobacterium nucleatum (22.5%). Streptococcus mutans (9.0%) and Actinomyces naeslundii (8.0%) were also frequently detected. Additionally, Prevotella intermedia (7.0%), Aggregatibacter actinomycetemcomitans (3.5%), and Enterococcus faecalis (2.5%) were present in lower frequencies. Conclusion: The presence of a diverse microbial flora in teeth with PELs underscores the polymicrobial nature of these lesions. The predominance of periodontal pathogens such as Porphyromonas gingivalis, Treponema denticola, and Fusobacterium nucleatum suggests a strong association between periodontal and endodontic infections. A comprehensive understanding of the microbial profile in PELs is crucial for tailored therapeutic approaches targeting the specific pathogens involved.

Mimicking hypomethylation of FUS requires liquid-liquid phase separation to induce synaptic dysfunctions.

The hypomethylation of fused in sarcoma (FUS) in frontotemporal lobar degeneration promotes the formation of irreversible condensates of FUS. However, the mechanisms by which these hypomethylated FUS condensates cause neuronal dysfunction are unknown. Here we report that expression of FUS constructs mimicking hypomethylated FUS causes aberrant dendritic FUS condensates in CA1 neurons. These hypomethylated FUS condensates exhibit spontaneous, and activity induced movement within the dendrite. They impair excitatory synaptic transmission, postsynaptic density-95 expression, and dendritic spine plasticity. These neurophysiological defects are dependent upon both the dendritic localisation of the condensates, and their ability to undergo liquid-liquid phase separation. These results indicate that the irreversible liquid-liquid phase separation is a key component of hypomethylated FUS pathophysiology in sporadic FTLD, and this can cause synapse dysfunction in sporadic FTLD.

The liquid-to-solid transition of FUS is promoted by the condensate surface.

A wide range of macromolecules can undergo phase separation, forming biomolecular condensates in living cells. These membraneless organelles are typically highly dynamic, formed reversibly, and carry out essential functions in biological systems. Crucially, however, a further liquid-to-solid transition of the condensates can lead to irreversible pathological aggregation and cellular dysfunction associated with the onset and development of neurodegenerative diseases. Despite the importance of this liquid-to-solid transition of proteins, the mechanism by which it is initiated in normally functional condensates is unknown. Here we show, by measuring the changes in structure, dynamics, and mechanics in time and space, that single-component FUS condensates do not uniformly convert to a solid gel, but rather that liquid and gel phases coexist simultaneously within the same condensate, resulting in highly inhomogeneous structures. Furthermore, our results show that this transition originates at the interface between the condensate and the dilute continuous phase, and once initiated, the gelation process propagates toward the center of the condensate. To probe such spatially inhomogeneous rheology during condensate aging, we use a combination of established micropipette aspiration experiments together with two optical techniques, spatial dynamic mapping and reflective confocal dynamic speckle microscopy. These results reveal the importance of the spatiotemporal dimension of the liquid-to-solid transition and highlight the interface of biomolecular condensates as a critical element in driving pathological protein aggregation.

Comparative Evaluation of Antimicrobial Efficacy of Calcium Hydroxide, Chlorhexidine, and Triple Antibiotic Paste in Different Combination Forms as Intracanal Medicaments against Enterococcus faecalis in Primary Teeth: An In Vivo Randomized Clinical Trial.

Aim: To compare and evaluate the antimicrobial efficacy against Enterococcus faecalis (E. faecalis) between a mix of calcium hydroxide [Ca(OH)2] powder and normal saline, a mix of Ca(OH)2 powder and 2% chlorhexidine (CHX) gluconate solution, a mix of triple antibiotic powder (TAP) and normal saline, and mix of TAP and 2% CHX gluconate solution. Materials and methods: A total of 60 teeth were included in the study. The first sample (S1) was collected after access opening from the widest canal of the tooth by inserting sterile absorbable paper point no 20 up to the full length of the canal for 1 minute. The second sample (S2) was collected after the chemomechanical preparation and irrigation. After that, subjects were randomly divided into four groups-group I-a mix of Ca(OH)2 and normal saline; group II-a mix of Ca(OH)2 and 2% CHX; group III-a mix of TAP and normal saline; and group IV-a mix of TAP and 2% CHX. Assigned intracanal medicaments were placed in the canals, and the teeth were temporarily sealed with a temporary restorative material. On the 7th day, canals were reopened and irrigated, and a third bacteriological sample (S3) was taken out. Later, canals were filled with suitable obturating material, followed by the placement of the permanent restoration. Results: There was a very highly significant (p < 0.005) difference in E. faecalis count in all the groups on day 7 after placement of intracanal medicament, being highest in group IV followed by group II, group III, and group I. Conclusion: Triple antibiotic powder (TAP) mixed with 2% CHX gluconate solution has superior antimicrobial efficacy against E. faecalis in primary teeth. How to cite this article: Qamar S, Jayanna R, Ahuja VR. Comparative Evaluation of Antimicrobial Efficacy of Calcium Hydroxide, Chlorhexidine, and Triple Antibiotic Paste in Different Combination Forms as Intracanal Medicaments against Enterococcus faecalis in Primary Teeth: An In Vivo Randomized Clinical Trial. Int J Clin Pediatr Dent 2023;16(3):448-452.

ANXA11 biomolecular condensates facilitate protein-lipid phase coupling on lysosomal membranes.

Phase transitions of cellular proteins and lipids play a key role in governing the organisation and coordination of intracellular biology. The frequent juxtaposition of proteinaceous biomolecular condensates to cellular membranes raises the intriguing prospect that phase transitions in proteins and lipids could be co-regulated. Here we investigate this possibility in the ribonucleoprotein (RNP) granule-ANXA11-lysosome ensemble, where ANXA11 tethers RNP granule condensates to lysosomal membranes to enable their co-trafficking. We show that changes to the protein phase state within this system, driven by the low complexity ANXA11 N-terminus, induce a coupled phase state change in the lipids of the underlying membrane. We identify the ANXA11 interacting proteins ALG2 and CALC as potent regulators of ANXA11-based phase coupling and demonstrate their influence on the nanomechanical properties of the ANXA11-lysosome ensemble and its capacity to engage RNP granules. The phenomenon of protein-lipid phase coupling we observe within this system offers an important template to understand the numerous other examples across the cell whereby biomolecular condensates closely juxtapose cell membranes.

Spatially non-uniform condensates emerge from dynamically arrested phase separation.

The formation of biomolecular condensates through phase separation from proteins and nucleic acids is emerging as a spatial organisational principle used broadly by living cells. Many such biomolecular condensates are not, however, homogeneous fluids, but possess an internal structure consisting of distinct sub-compartments with different compositions. Notably, condensates can contain compartments that are depleted in the biopolymers that make up the condensate. Here, we show that such double-emulsion condensates emerge via dynamically arrested phase transitions. The combination of a change in composition coupled with a slow response to this change can lead to the nucleation of biopolymer-poor droplets within the polymer-rich condensate phase. Our findings demonstrate that condensates with a complex internal architecture can arise from kinetic, rather than purely thermodynamic driving forces, and provide more generally an avenue to understand and control the internal structure of condensates in vitro and in vivo.

Biomolecular condensate phase diagrams with a combinatorial microdroplet platform.

The assembly of biomolecules into condensates is a fundamental process underlying the organisation of the intracellular space and the regulation of many cellular functions. Mapping and characterising phase behaviour of biomolecules is essential to understand the mechanisms of condensate assembly, and to develop therapeutic strategies targeting biomolecular condensate systems. A central concept for characterising phase-separating systems is the phase diagram. Phase diagrams are typically built from numerous individual measurements sampling different parts of the parameter space. However, even when performed in microwell plate format, this process is slow, low throughput and requires significant sample consumption. To address this challenge, we present here a combinatorial droplet microfluidic platform, termed PhaseScan, for rapid and high-resolution acquisition of multidimensional biomolecular phase diagrams. Using this platform, we characterise the phase behaviour of a wide range of systems under a variety of conditions and demonstrate that this approach allows the quantitative characterisation of the effect of small molecules on biomolecular phase transitions.

Mitochondrial ROS control neuronal excitability and cell fate in frontotemporal dementia.

INTRODUCTION: The second most common form of early-onset dementia-frontotemporal dementia (FTD)-is often characterized by the aggregation of the microtubule-associated protein tau. Here we studied the mechanism of tau-induced neuronal dysfunction in neurons with the FTD-related 10+16 MAPT mutation. METHODS: Live imaging, electrophysiology, and redox proteomics were used in 10+16 induced pluripotent stem cell-derived neurons and a model of tau spreading in primary cultures. RESULTS: Overproduction of mitochondrial reactive oxygen species (ROS) in 10+16 neurons alters the trafficking of specific glutamate receptor subunits via redox regulation. Increased surface expression of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptors containing GluA1 and NR2B subunits leads to impaired glutamatergic signaling, calcium overload, and excitotoxicity. Mitochondrial antioxidants restore the altered response and prevent neuronal death. Importantly, extracellular 4R tau induces the same pathological response in healthy neurons, thus proposing a mechanism for disease propagation. DISCUSSION: These results demonstrate mitochondrial ROS modulate glutamatergic signaling in FTD, and suggest a new therapeutic strategy.

Differential interaction with TREM2 modulates microglial uptake of modified Aß species.

Rare coding variants of the microglial triggering receptor expressed on myeloid cells 2 (TREM2) confer an increased risk for Alzheimer's disease (AD) characterized by the progressive accumulation of aggregated forms of amyloid ß peptides (Aß). Aß peptides are generated by proteolytic processing of the amyloid precursor protein (APP). Heterogeneity in proteolytic cleavages and additional post-translational modifications result in the production of several distinct Aß variants that could differ in their aggregation behavior and toxic properties. Here, we sought to assess whether post-translational modifications of Aß affect the interaction with TREM2. Biophysical and biochemical methods revealed that TREM2 preferentially interacts with oligomeric Aß, and that phosphorylation of Aß increases this interaction. Phosphorylation of Aß also affected the TREM2 dependent interaction and phagocytosis by primary microglia and in APP transgenic mouse models. Thus, TREM2 function is important for sensing phosphorylated Aß variants in distinct aggregation states and reduces the accumulation and deposition of these toxic Aß species in preclinical models of Alzheimer's disease.

Massive gingival enlargement in a nine-year-old paediatric patient: A rare case report.

Gingival enlargement (GE) is a well-known clinical phenomena with the primary aetiology being plaque and poor oral hygiene. Many reasons for GE have been known. Most of the time good oral hygiene is sufficient to achieve normal healthy gingiva. GE is a heterogeneous group of disorder characterized by progressive enlargement of the gingiva with an increase in submucosal connective tissue elements. Some of them are inherited and iatrogenic while others are idiopathic. In this case, we report a case with massive idiopathic GE in a 9-year-old female child; treatment received, histopathological description and follow-up are discussed.

Wild-type sTREM2 blocks Aß aggregation and neurotoxicity, but the Alzheimer's R47H mutant increases Aß aggregation.

TREM2 is a pattern recognition receptor, expressed on microglia and myeloid cells, detecting lipids and Aß and inducing an innate immune response. Missense mutations (e.g., R47H) of TREM2 increase risk of Alzheimer's disease (AD). The soluble ectodomain of wild-type TREM2 (sTREM2) has been shown to protect against AD in vivo, but the underlying mechanisms are unclear. We show that Aß oligomers bind to cellular TREM2, inducing shedding of the sTREM2 domain. Wild-type sTREM2 bound to Aß oligomers (measured by single-molecule imaging, dot blots, and Bio-Layer Interferometry) inhibited Aß oligomerization and disaggregated preformed Aß oligomers and protofibrils (measured by transmission electron microscopy, dot blots, and size-exclusion chromatography). Wild-type sTREM2 also inhibited Aß fibrillization (measured by imaging and thioflavin T fluorescence) and blocked Aß-induced neurotoxicity (measured by permeabilization of artificial membranes and by loss of neurons in primary neuronal-glial cocultures). In contrast, the R47H AD-risk variant of sTREM2 is less able to bind and disaggregate oligomeric Aß but rather promotes Aß protofibril formation and neurotoxicity. Thus, in addition to inducing an immune response, wild-type TREM2 may protect against amyloid pathology by the Aß-induced release of sTREM2, which blocks Aß aggregation and neurotoxicity. In contrast, R47H sTREM2 promotes Aß aggregation into protofibril that may be toxic to neurons. These findings may explain how wild-type sTREM2 apparently protects against AD in vivo and why a single copy of the R47H variant gene is associated with increased AD risk.

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions.

Liquid-liquid phase separation of proteins underpins the formation of membraneless compartments in living cells. Elucidating the molecular driving forces underlying protein phase transitions is therefore a key objective for understanding biological function and malfunction. Here we show that cellular proteins, which form condensates at low salt concentrations, including FUS, TDP-43, Brd4, Sox2, and Annexin A11, can reenter a phase-separated regime at high salt concentrations. By bringing together experiments and simulations, we demonstrate that this reentrant phase transition in the high-salt regime is driven by hydrophobic and non-ionic interactions, and is mechanistically distinct from the low-salt regime, where condensates are additionally stabilized by electrostatic forces. Our work thus sheds light on the cooperation of hydrophobic and non-ionic interactions as general driving forces in the condensation process, with important implications for aberrant function, druggability, and material properties of biomolecular condensates.

Effectiveness of Platelet-Rich Fibrin in the Treatment of Intrabony Defects with or Without Bone Graft: A Clinical Comparative Study.

BACKGROUND: Periodontal diseases are a group of inflammatory diseases causing alveolar bone loss and eventually leading to loss of teeth. The present study was evaluated the effectiveness of platelet-rich fibrin (PRF) in the treatment of intrabony defects with or without bone graft. MATERIALS AND METHODS: Thirty subjects with the presence of intrabony defects were enrolled. All subjects were included irrespective of age and gender. A questionnaire was prepared for extracting demographic and personal details of all the patients. William probe and moth mirror-tweezers set was used for carrying out clinical examination of all subjects. Random and unbiased division of all the subjects was done with ten patients in each group as follows: Group I: Subjects in which treatment was carried out using PRF with demineralized bone matrix, Group II: Subjects in which treatment was carried out using PRF alone, and Group III: Subjects in which treatment was carried in the form of open flap debridement (OFD). Pretreatment and posttreatment clinical variables were assessed which included plaque index (PI), gingival index (GI), probing depth (PD), relative attachment level (RAL), and gingival recession (GR) were assessed at baseline and 9 months postoperatively were calculated. RESULTS: Mean PI among Group 1, Group 2, and Group 3 at baseline was 0.78, 0.8, and 0.84, respectively. Mean PI among Group 1, Group 2, and Group 3 at 9 months follow-up was 0.56, 0.55, and 0.72, respectively. Significant results were obtained while comparing the PI among the three study groups at follow-up. Mean GI among Group 1, Group 2, and Group 3 at baseline was 0.78, 0.8, and 0.84, respectively. Mean GI among Group 1, Group 2, and Group 3 at 9 months follow-up was 0.56, 0.55, and 0.72, respectively. Significant results were obtained while comparing the GI among the three study groups at follow-up. Significant difference was seen in PD, RAL, and GR from baseline to 9 months in all groups (P < 0.05). CONCLUSION: PRF leads to significantly better improvement in the clinical parameters on follow-up in comparison to OFD alone in patients with the presence of intrabony defects.

Biomolecular condensates undergo a generic shear-mediated liquid-to-solid transition.

Membrane-less organelles resulting from liquid-liquid phase separation of biopolymers into intracellular condensates control essential biological functions, including messenger RNA processing, cell signalling and embryogenesis1-4. It has recently been discovered that several such protein condensates can undergo a further irreversible phase transition, forming solid nanoscale aggregates associated with neurodegenerative disease5-7. While the irreversible gelation of protein condensates is generally related to malfunction and disease, one case where the liquid-to-solid transition of protein condensates is functional, however, is that of silk spinning8,9. The formation of silk fibrils is largely driven by shear, yet it is not known what factors control the pathological gelation of functional condensates. Here we demonstrate that four proteins and one peptide system, with no function associated with fibre formation, have a strong propensity to undergo a liquid-to-solid transition when exposed to even low levels of mechanical shear once present in their liquid-liquid phase separated form. Using microfluidics to control the application of shear, we generated fibres from single-protein condensates and characterized their structural and material properties as a function of shear stress. Our results reveal generic backbone-backbone hydrogen bonding constraints as a determining factor in governing this transition. These observations suggest that shear can play an important role in the irreversible liquid-to-solid transition of protein condensates, shed light on the role of physical factors in driving this transition in protein aggregation-related diseases and open a new route towards artificial shear responsive biomaterials.

Role of natural salivary defenses in the maintenance of healthy oral microbiota in children and adolescents.

AIM: The present study served the purpose of assessing the levels of salivary immunoglobulin A (IgA), immunoglobulin G (IgG), proteins, calcium, inorganic phosphorus, and alkaline phosphatase levels in caries-free and caries active children. MATERIALS AND METHODS: Stratified randomized sampling method was used to include 40 subjects in the age group 12-15 years having a full complement of permanent dentition except for third molars. The selected pediatric subjects were further divided into two groups of 20 each based on DMFS score, Group-I - Caries free (DMFS score = 0) and Group-II - Caries active (DMFS score ≥10). Unstimulated midmorning saliva samples were collected and analyzed colorimetrically and by radial immunodiffusion method for constituents of saliva understudy. RESULTS: The mean salivary IgA levels in children in Group-I (caries-free children) was 10.63 ± 2.85 mg/dL which was statistically higher as compared to caries active children in Group-II (8.50 ± 1.43 mg/dL).The mean salivary protein level in children of Group-II was statistically higher at 3.28 ± 0.12 mg/dL as compared to Group-I (2.89 ± 0.11 mg/dL). CONCLUSION: The present study showed decreased levels of salivary immunoglobulin A and high concentration of salivary protein in children with increased caries experience which is indicative of the protective role of salivary constituents in caries-free children.

A Comparative Study of Platelet-rich Fibrin (PRF) and Titanium-prepared Platelet-rich Fibrin (T-PRF) in Management of Endo-perio Lesions.

AIM: The aim of the study was to compare the platelet-rich fibrin (PRF) and titanium-prepared platelet-rich fibrin (T-PRF) in the management of endo-perio lesions. MATERIALS AND METHODS: This study was conducted with 140 patients who are affected by endo-perio lesions, and the patients were divided into two groups. In group I, patients were treated with PRF, and in group II, patients were treated with T-PRF. Endodontic treatment was done in all cases, following a standardized aseptic method. Probing pocket and relative attachment level were recorded after 3 months and 6 months in both groups. RESULTS: After 3 months, the mean change of probing pocket depth (PPD) was 68% in group I and 33.41% in group II. The intergroup distinction was not significant (p > 0.05), whereas the intragroup evaluation was significant (p < 0.05). The mean change after 6 months was 42.59% in group I and 43.90% in group II. The difference was not significant (p > 0.05). After 3 months, the mean% change of relative attachment level (RAL) was 31.20% in group I and 31.60% in group II. The intergroup distinction was not significant (p > 0.05), whereas the intragroup evaluation established a considerable discrepancy (p < 0.05). The mean change after 6 months was 40.82% in group I and 42.12% in group II. CONCLUSION: Both PRF and T-PRF were effective in inducing a reduction in pocket depth and useful in gaining attachment level. CLINICAL SIGNIFICANCE: With the use of PRF and T-PRF, the complex case of endo-perio lesions can be efficiently managed.

Inherited and Sporadic Amyotrophic Lateral Sclerosis and Fronto-Temporal Lobar Degenerations arising from Pathological Condensates of Phase Separating Proteins.

Recent work on the biophysics of proteins with low complexity, intrinsically disordered domains that have the capacity to form biological condensates has profoundly altered the concepts about the pathogenesis of inherited and sporadic neurodegenerative disorders associated with pathological accumulation of these proteins. In the present review, we use the FUS, TDP-43 and A11 proteins as examples to illustrate how missense mutations and aberrant post-translational modifications of these proteins cause amyotrophic lateral sclerosis (ALS) and fronto-temporal lobar degeneration (FTLD).

RNA Granules Hitchhike on Lysosomes for Long-Distance Transport, Using Annexin A11 as a Molecular Tether.

Long-distance RNA transport enables local protein synthesis at metabolically-active sites distant from the nucleus. This process ensures an appropriate spatial organization of proteins, vital to polarized cells such as neurons. Here, we present a mechanism for RNA transport in which RNA granules "hitchhike" on moving lysosomes. In vitro biophysical modeling, live-cell microscopy, and unbiased proximity labeling proteomics reveal that annexin A11 (ANXA11), an RNA granule-associated phosphoinositide-binding protein, acts as a molecular tether between RNA granules and lysosomes. ANXA11 possesses an N-terminal low complexity domain, facilitating its phase separation into membraneless RNA granules, and a C-terminal membrane binding domain, enabling interactions with lysosomes. RNA granule transport requires ANXA11, and amyotrophic lateral sclerosis (ALS)-associated mutations in ANXA11 impair RNA granule transport by disrupting their interactions with lysosomes. Thus, ANXA11 mediates neuronal RNA transport by tethering RNA granules to actively-transported lysosomes, performing a critical cellular function that is disrupted in ALS.

Oligomer Diversity during the Aggregation of the Repeat Region of Tau.

The molecular mechanism of protein aggregation is of both fundamental and clinical importance as amyloid aggregates are linked to a number of neurodegenerative disorders. Such protein aggregates include macroscopic insoluble fibrils as well as small soluble oligomeric species. Time-dependent resolution of these species is prerequisite for a detailed quantitative understanding of protein aggregation; this remains challenging due to the lack of methods for detecting and characterizing transient and heterogeneous protein oligomers. Here we have used single molecule fluorescence techniques combined with mechanistic modeling to study the heparin-induced aggregation of the repeat region of tau, which forms the core region of neurofibrillary tangles found in Alzheimer's disease. We distinguish several subpopulations of oligomers with different stability and follow their evolution during aggregation reactions as a function of temperature and concentration. Employment of techniques from chemical kinetics reveals that the two largest populations are structurally distinct from fibrils and are both kinetically and thermodynamically unstable. The first population is in rapid exchange with monomers and held together by electrostatic interactions; the second is kinetically more stable, dominates at later times, and is probably off-pathway to fibril formation. These more stable oligomers may contribute to other oligomer induced effects in the cellular environment, for example, by overloading protein quality control systems. We also show that the shortest growing filaments remain suspended in aqueous buffer and thus comprise a third, smaller population of transient oligomers with cross-ß structure. Overall our data show that a diverse population of oligomers of different structures and half-lives are formed during the aggregation reaction with the great majority of oligomers formed not going on to form fibrils.