Low-complexity Regions Research Articles

Air Traffic Complexity Map Based on Linear Dynamical Systems

This paper presents a new air traffic complexity metric based on linear dynamical systems, of which the goal is to quantify the intrinsic complexity of a set of aircraft trajectories. Previous works have demonstrated that the structure and organization of air traffic are essential factors in the perception of the complexity of an air traffic situation. Usually, they were not able to explicitly address trajectory pattern organization. The new metric, by identifying the organization properties of trajectories in a traffic pattern, captures some of the key factors involved in ATC complexity. The key idea of this work is to find a linear dynamical system which fits a vector field as closely as possible to the observations given by the aircraft positions and speeds. This approach produces an aggregate complexity metric that enables one to identify high (low) complexity regions of the airspace and compare their relative complexity. The metric is very appropriate to compare different traffic situations for any scale (sector or country) by associating a complexity index to each trajectory sample in the airspace. For instance, to compute the complexity for a sector, one must just sum-up the complexity for trajectory samples intersecting such a sector. This computation can also be extended in the time dimension in order to estimate the average complexity in a given airspace for a period of time.

Gel‐like inclusions of C‐terminal fragments of TDP‐43 sequester stalled proteasomes in neurons

Aggregation of the multifunctional RNA‐binding protein TDP‐43 defines large subgroups of amyotrophic lateral sclerosis and frontotemporal dementia and correlates with neurodegeneration in both diseases. In disease, characteristic C‐terminal fragments of ~25 kDa ("TDP‐25") accumulate in cytoplasmic inclusions. Here, we analyze gain‐of‐function mechanisms of TDP‐25 combining cryo‐electron tomography, proteomics, and functional assays. In neurons, cytoplasmic TDP‐25 inclusions are amorphous, and photobleaching experiments reveal gel‐like biophysical properties that are less dynamic than nuclear TDP‐43. Compared with full‐length TDP‐43, the TDP‐25 interactome is depleted of low‐complexity domain proteins. TDP‐25 inclusions are enriched in 26S proteasomes adopting exclusively substrate‐processing conformations, suggesting that inclusions sequester proteasomes, which are largely stalled and no longer undergo the cyclic conformational changes required for proteolytic activity. Reporter assays confirm that TDP‐25 impairs proteostasis, and this inhibitory function is enhanced by ALS‐causing TDP‐43 mutations. These findings support a patho‐physiological relevance of proteasome dysfunction in ALS/FTD.

Investigating the disordered regions (MoRFs, SLiMs and LCRs) and functions of mimicry proteins/peptides in silico

Microbial mimicry of the host proteins/peptides can elicit host auto-reactive T- or B-cells resulting in autoimmune disease(s). Since intrinsically disordered protein regions (IDPRs) are involved in several host cell signaling and PPI networks, molecular mimicry of the IDPRs can help the pathogens in substituting their own proteins in the host cell-signaling and PPI networks and, ultimately hijacking the host cellular machinery. Thus, the present study was conducted to discern the structural disorder and intrinsically disordered protein regions (IDPRs) like, molecular recognition features (MoRFs), short linear motifs (SLiMs), and low complexity regions (LCRs) in the experimentally verified mimicry proteins and peptides (mimitopes) of bacteria, viruses and host. Also, functional characteristics of the mimicry proteins were studied in silico. Our results indicated that 78% of the bacterial host mimicry proteins and 45% of the bacterial host mimitopes were moderately/highly disordered while, 73% of the viral host mimicry proteins and 31% of the viral host mimitopes were moderately/highly disordered. Among the pathogens, 27% of the bacterial mimicry proteins and 13% of the bacterial mimitopes were moderately/highly disordered while, 53% of the viral mimicry proteins and 21% of the viral mimitopes were moderately/highly disordered. Though IDPR were frequent in host, bacterial and viral mimicry proteins, only a few mimitopes overlapped with the IDPRs like, MoRFs, SLiMs and LCRs. This suggests that most of the microbes cannot use molecular mimicry to modulate the host PPIs and hijack the host cell machinery. Functional analyses indicated that most of the pathogens exhibited mimicry with the host proteins involved in ion binding and signaling pathways. This is the first report on the disordered regions and functional aspects of experimentally proven host and microbial mimicry proteins.

Dual-gRNA approach with limited off-target effect corrects C9ORF72 repeat expansion in vivo

C9ORF72 GGGGCC repeat expansion is the most common genetic cause for amyotrophic lateral sclerosis and frontotemporal dementia, which generates abnormal DNA and RNA structures and produces toxic proteins. Recently, efficacy of CRISPR/Cas9-mediated editing has been proven in treatment of disease. However, DNA low complexity surrounding C9ORF72 expansion increases the off-target risks. Here we provide a dual-gRNA design outside of the low complexity region which enables us to remove the repeat DNA in a ‘cutting-deletion-fusion’ manner with a high fusion efficiency (50%). Our dual-gRNA design limits off-target effect and does not significantly affect C9ORF72 expression. In neurons carrying patient C9ORF72 expansion, our approach removes the repeat DNA and corrects the RNA foci in vitro and in vivo. Therefore, we conclude that our proof-of-concept design correct C9ORF72 repeat expansion, which may have potential therapeutic value for the patients.

Full-Length Genome of an Ogataea polymorpha Strain CBS4732 ura3Δ Reveals Large Duplicated Segments in Subtelomeric Regions

BackgroundCurrently, methylotrophic yeasts (e.g., Pichia pastoris, Ogataea polymorpha, and Candida boindii) are subjects of intense genomics studies in basic research and industrial applications. In the genus Ogataea, most research is focused on three basic O. polymorpha strains-CBS4732, NCYC495, and DL-1. However, the relationship between CBS4732, NCYC495, and DL-1 remains unclear, as the genomic differences between them have not be exactly determined without their high-quality complete genomes. As a nutritionally deficient mutant derived from CBS4732, the O. polymorpha strain CBS4732 ura3Δ (named HU-11) is being used for high-yield production of several important proteins or peptides. HU-11 has the same reference genome as CBS4732 (noted as HU-11/CBS4732), because the only genomic difference between them is a 5-bp insertion.ResultsIn the present study, we have assembled the full-length genome of O. polymorpha HU-11/CBS4732 using high-depth PacBio and Illumina data. Long terminal repeat retrotransposons (LTR-rts), rDNA, 5′ and 3′ telomeric, subtelomeric, low complexity and other repeat regions were exactly determined to improve the genome quality. In brief, the main findings include complete rDNAs, complete LTR-rts, three large duplicated segments in subtelomeric regions and three structural variations between the HU-11/CBS4732 and NCYC495 genomes. These findings are very important for the assembly of full-length genomes of yeast and the correction of assembly errors in the published genomes of Ogataea spp. HU-11/CBS4732 is so phylogenetically close to NCYC495 that the syntenic regions cover nearly 100% of their genomes. Moreover, HU-11/CBS4732 and NCYC495 share a nucleotide identity of 99.5% through their whole genomes. CBS4732 and NCYC495 can be regarded as the same strain in basic research and industrial applications.ConclusionThe present study preliminarily revealed the relationship between CBS4732, NCYC495, and DL-1. Our findings provide new opportunities for in-depth understanding of genome evolution in methylotrophic yeasts and lay the foundations for the industrial applications of O. polymorpha CBS4732, NCYC495, DL-1, and their derivative strains. The full-length genome of O. polymorpha HU-11/CBS4732 should be included into the NCBI RefSeq database for future studies of Ogataea spp.

On the development of a neoantigen vaccine for the prevention of Lynch Syndrome

Lynch Syndrome (LS) is an autosomal dominant genetic condition that causes a high risk of colorectal cancer. The hallmark of LS is genetic instability as a result of mismatch repair (MMR) deficiency, particularly in repetitive low complexity regions called microsatellites (MS). MLH1-/- mice deficient in MMR are prone to developing tumors in the colon, upon oral administration of dextran sodium sulfate (DSS), at a rate of more than 70%. Using this LS mouse model, we found a novel tumor neo-antigen from a deletion mutation of the coding MS in the SENP6 gene that prevented tumorigenesis or hindered tumor growth rate in immunized mice. This was accomplished via high throughput exome sequencing of DSS-induced colorectal tumors in the MLH1-/- mice and predicting the most highly immunogenic mutant gene products processed and presented as antigens in C57BL/6 MHC-I molecules. Throughout our study, we were able to prove the validity of the vaccine by analyzing the colorectal tumors in immunized DSS-treated mice using either our epitope, called Sp6D1, or an unrelated peptide as a negative control. Tumors developed in this context were found to be antigenic and Sp6D1-specific CD8+ tumor infiltrating lymphocytes were detected by flow cytometry and cytotoxic T lymphocytes (CTL) killing assays. Additionally, immunohistochemistry showed that tumor-adjacent tertiary lymphoid organs were a potentially significant source of CD8+ lymphocytes. Altogether, our results indicate that there may be a protective effect to patients carrying LS mutations through the induction of a peptide-specific CTL response from the use of neoepitope vaccination.

The role of ATP in solubilizing RNA-binding protein fused in sarcoma.

Intrinsically disordered protein (IDP) plays an important role in liquid-liquid phase separation (LLPS). RNA-binding protein fused in sarcoma (FUS) is a well-studied IDP that induces LLPS since its low-complexity core region (FUS-LC-core) is essential for droplet formation through contacts between FUS-LC-cores. Several experimental studies have reported that adenosine triphosphate (ATP) concentrations modulate LLPS-driven droplet formation through the dissolution of FUS. To elucidate the role of ATP in this dissolution, microsecond-order all-atom molecular dynamics (MD) simulations were performed for a crowded system of FUS-LC-cores in the presence of multiple ATP molecules. Our analysis revealed that the adenine group of ATP frequently contacted the FUS-LC-core, and the phosphoric acid group of ATP was exposed to the external solvent, which promoted both hydration and solubilization of FUS.

Establishing analytical validity of BeadChip array genotype data by comparison to whole-genome sequence and standard benchmark datasets

BackgroundClinical use of genotype data requires high positive predictive value (PPV) and thorough understanding of the genotyping platform characteristics. BeadChip arrays, such as the Global Screening Array (GSA), potentially offer a high-throughput, low-cost clinical screen for known variants. We hypothesize that quality assessment and comparison to whole-genome sequence and benchmark data establish the analytical validity of GSA genotyping.MethodsTo test this hypothesis, we selected 263 samples from Coriell, generated GSA genotypes in triplicate, generated whole genome sequence (rWGS) genotypes, assessed the quality of each set of genotypes, and compared each set of genotypes to each other and to the 1000 Genomes Phase 3 (1KG) genotypes, a performance benchmark. For 59 genes (MAP59), we also performed theoretical and empirical evaluation of variants deemed medically actionable predispositions.ResultsQuality analyses detected sample contamination and increased assay failure along the chip margins. Comparison to benchmark data demonstrated that > 82% of the GSA assays had a PPV of 1. GSA assays targeting transitions, genomic regions of high complexity, and common variants performed better than those targeting transversions, regions of low complexity, and rare variants. Comparison of GSA data to rWGS and 1KG data showed > 99% performance across all measured parameters. Consistent with predictions from prior studies, the GSA detection of variation within the MAP59 genes was 3/261.ConclusionWe establish the analytical validity of GSA assays using quality analytics and comparison to benchmark and rWGS data. GSA assays meet the standards of a clinical screen although assays interrogating rare variants, transversions, and variants within low-complexity regions require careful evaluation.

Proteomic analysis of heat-stable proteins revealed an increased proportion of proteins with compositionally biased regions

Intrinsically disordered proteins (IDPs) have been in the spotlight for their unique properties, such as their lack of secondary structures and low sequence complexity. Alpha-synuclein and tau are representative disease-related IDPs with low complexity regions in their sequences, accumulating in the brains of patients with Parkinson disease and Alzheimer disease, respectively. Their heat resistance in particular was what attracted our attention. We assumed that there exist many other unidentified proteins that are resistant to heat-treatment, referred to as heat-stable proteins, which would also have low sequence complexity. In this study, we performed proteomic analysis of heat-stable proteins of mouse brains and found that proteins with compositionally biased regions are abundant in the heat-stable proteins. The proteins related to neurodegeneration are known to undergo different types of post-translational modifications (PTMs) such as phosphorylation and ubiquitination. We then investigated the heat-stability and aggregation properties of phosphorylated synuclein and tau with different phosphorylation sites. We suggest that PTMs can be important factors that determine the heat-stability and aggregation properties of a protein. IDPs identified in the heat-stable proteins of mouse brains would be candidates for the pathogenic proteins for neurodegeneration.

EP343: Long-read genome sequencing informs the molecular etiology of imprinting disorders

Although the majority of rare diseases are suspected to be genetic in origin, the causal genes are often unknown. Consequently, families affected by rare diseases go through challenging diagnostic odysseys that involve multiple care providers, serial testing, and span several years. Exome and genome sequencing (GS) have improved rates of diagnosis, allowing agnostic characterization of a broad spectrum of genetic variation. Recent long-read sequencing (LRS) has overcome some of the limitations of short-read sequencing for low complexity and repetitive regions of the genome.

Two short low complexity regions (LCRs) are hallmark sequences of the Delta SARS-CoV-2 variant spike protein

Low complexity regions (LCRs) are protein sequences formed by a set of compositionally biased residues. LCRs are extremely abundant in cellular proteins and have also been reported in viruses, where they may partake in evasion of the host immune system. Analyses of 28,231 SARS-CoV-2 whole proteomes and of 261,051 spike protein sequences revealed the presence of four extremely conserved LCRs in the spike protein of several SARS-CoV-2 variants. With the exception of Iota, where it is absent, the Spike LCR-1 is present in the signal peptide of 80.57% of the Delta variant sequences, and in other variants of concern and interest. The Spike LCR-2 is highly prevalent (79.87%) in Iota. Two distinctive LCRs are present in the Delta spike protein. The Delta Spike LCR-3 is present in 99.19% of the analyzed sequences, and the Delta Spike LCR-4 in 98.3% of the same set of proteins. These two LCRs are located in the furin cleavage site and HR1 domain, respectively, and may be considered hallmark traits of the Delta variant. The presence of the medically-important point mutations P681R and D950N in these LCRs, combined with the ubiquity of these regions in the highly contagious Delta variant opens the possibility that they may play a role in its rapid spread.

Using synthetic chromosome controls to evaluate the sequencing of difficult regions within the human genome

BackgroundNext-generation sequencing (NGS) can identify mutations in the human genome that cause disease and has been widely adopted in clinical diagnosis. However, the human genome contains many polymorphic, low-complexity, and repetitive regions that are difficult to sequence and analyze. Despite their difficulty, these regions include many clinically important sequences that can inform the treatment of human diseases and improve the diagnostic yield of NGS.ResultsTo evaluate the accuracy by which these difficult regions are analyzed with NGS, we built an in silico decoy chromosome, along with corresponding synthetic DNA reference controls, that encode difficult and clinically important human genome regions, including repeats, microsatellites, HLA genes, and immune receptors. These controls provide a known ground-truth reference against which to measure the performance of diverse sequencing technologies, reagents, and bioinformatic tools. Using this approach, we provide a comprehensive evaluation of short- and long-read sequencing instruments, library preparation methods, and software tools and identify the errors and systematic bias that confound our resolution of these remaining difficult regions.ConclusionsThis study provides an analytical validation of diagnosis using NGS in difficult regions of the human genome and highlights the challenges that remain to resolve these difficult regions.

Regions with two amino acids in protein sequences: A step forward from homorepeats into the low complexity landscape

Low complexity regions (LCRs) differ in amino acid composition from the background provided by the corresponding proteomes. The simplest LCRs are homorepeats (or polyX), regions composed of mostly-one amino acid type. Extensive research has been done to characterize homorepeats, and their taxonomic, functional and structural features depend on the amino acid type and sequence context. From them, the next step towards the study of LCRs are the regions composed of two types of amino acids, which we call polyXY. We classify polyXY in three categories based on the arrangement of the two amino acid types ‘X’ and ‘Y’: direpeats (e.g. ‘XYXYXY’), joined (e.g. ‘XXXYYY’) and shuffled (e.g. ‘XYYXXY’). We developed a script to search for polyXY, and located them in a comprehensive set of 20,340 reference proteomes. These results are available in a dedicated web server called XYs, in which the user can also submit their own protein datasets to detect polyXY. We studied the distribution of polyXY types by amino acid pair XY and category, and show that polyXY in Eukaryota are mainly located within intrinsically disordered regions. Our study provides a first step towards the characterization of polyXY as protein motifs.

Glycogen Synthase Kinase-3 Isoform Variants and Their Inhibitory Phosphorylation in Human Testes and Spermatozoa.

To clarify (phospho-) glycogen synthase kinase-3 (GSK3) isoform variants in the germline and soma of human testes and spermatozoa. GSK3 isoform variants in normospermatogenic and Sertoli cell-only (SCO) testicular biopsies and spermatozoa were examined. In normospermatogenic testes, GSK3α and GSK3β variants 1 and 2 different in low complexity region (LCR) were expressed and their levels were decreased in SCO testes. GSK3β variant 3 was only expressed in SCO testes. GSK3β as well as GSK3α, the dominant isoforms in testes were decreased in SCO testes. In normospermatogenic testes, GSK3β were found in spermatogonia and markedly decreased in meiotic germ cells in which GSK3α was dominant. p-GSK3α/β were marginal in spermatogonia and early spermatocytes. In SCO testes, GSK3α/β immunoreactivity in seminiferous epithelia was weaker than those of normospermatogenic testes whereas p-GSK3α/β(Ser) immunoreactivity was visibly increased in Sertoli cells. GSK3α was dominant in ejaculated spermatozoa in which GSK3α and p-GSK3α(Ser) were found in the head, midpiece, and tail. In acrosome-reacted spermatozoa, GSK3α was found in the equatorial region of head, midpiece, and tail, and p-GSK3α(Ser) was only found in midpiece. During sperm capacitation, p-GSK3α(Ser) was significantly increased together with phosphotyrosine proteins and motility. In human male germ cells, GSK3 isoforms different in LCRs switch from GSK3β to GSK3α during meiotic entry, suggesting the isoform-specific roles of GSK3α and GSK3β in meiosis and stemness or proliferation of spermatogonia, respectively. In dormant Sertoli cells of SCO testes kinase activity of GSK3 might be downregulated via inhibitory phosphorylation. In spermatozoa, inhibitory phosphorylation of GSK3α might be coupled with activation of motility during capacitation.

RNF219 attenuates global mRNA decay through inhibition of CCR4-NOT complex-mediated deadenylation

The CCR4-NOT complex acts as a central player in the control of mRNA turnover and mediates accelerated mRNA degradation upon HDAC inhibition. Here, we explored acetylation-induced changes in the composition of the CCR4-NOT complex by purification of the endogenously tagged scaffold subunit NOT1 and identified RNF219 as an acetylation-regulated cofactor. We demonstrate that RNF219 is an active RING-type E3 ligase which stably associates with CCR4-NOT via NOT9 through a short linear motif (SLiM) embedded within the C-terminal low-complexity region of RNF219. By using a reconstituted six-subunit human CCR4-NOT complex, we demonstrate that RNF219 inhibits deadenylation through the direct interaction of the α-helical SLiM with the NOT9 module. Transcriptome-wide mRNA half-life measurements reveal that RNF219 attenuates global mRNA turnover in cells, with differential requirement of its RING domain. Our results establish RNF219 as an inhibitor of CCR4-NOT-mediated deadenylation, whose loss upon HDAC inhibition contributes to accelerated mRNA turnover.

Bioinspired translation of classical music into de novo protein structures using deep learning and molecular modeling

Architected biomaterials, as well as sound and music, are constructed from small building blocks that are assembled across time- and length-scales. Here we present a novel deep learning-enabled integrated algorithmic workflow to merge the two concepts for radical discovery of de novo protein materials, exploiting musical creativity as the foundation, and extrapolating through a recursive method to increase protein complexity by successively injecting protein chemistry into the process. Indeed, music is one of the few universal expressions that can create bridges between cultures, find associations between seemingly unrelated concepts, and can be used as a novel way to generate bio-inspired designs that derive functions from the imaginations of the creative mind. Earlier work has offered a pathway to convert proteins into sound, and sound into proteins. Here we build on this paradigm and translate a piece of classical music into matter. Based on Bach’s Goldberg variations, we offer a series of case studies to convert the musical data imagined by the composer into protein design, and folded into a 3D structure using deep learning. The quest we seek to address is to identify semblances, or memories, or information content in such musical creation, that offers new insights into pattern relationships between distinct manifestations of information. Using basic local alignment search tool analysis, we find that several fragments of the new proteins display similarities to existing protein sequences found in proteobacteria among other organisms, especially in regions of low complexity and repetitive motifs. The resulting protein forms the basis for iterative musical composition, and an evolutionary paradigm that defines a variational pathway for melodic development, complementing conventional creative or mathematical methods. This paper broadens the concept of what is understood as bio-inspiration to include a broad array of systems created by humans, animals, or other natural mechanisms.

Genomic Subgroups Impact Post-Transplant Survival in Patients with Myelodysplastic Syndrome: A CIBMTR Analysis

BackgroundMyelodysplastic syndromes (MDS) are clonal stem cell malignancies characterized by cytopenia, inefficient hematopoiesis, dysplasia in one or more myeloid cell lineages and increased risk of development of acute myeloid leukemia. It has been well appreciated that genomic alterations play a key role in MDS pathogenesis. The Revised International Prognostic Scoring System (IPSS-R) algorithm is commonly used to predict overall survival but may fail to recapitulate reliable prognostic information at the individual patient level, especially at the time of hematopoietic cell transplantation (HCT). Current World Health Organization (WHO) classification includes MDS with isolated 5q deletion as the only genetically defined category. Comprehensive analysis of recurrent genomic features by unsupervised clustering empowers discovery of potential prognostic molecular signatures.MethodsUsing whole blood samples obtained from 494 MDS patients at the time of HCT, we conducted whole-genome sequencing (WGS) and somatic variant processing via a custom analytic pipeline based on OCTOPUS and a set of annotation databases. Multiple filters allowed for selection and fine-tuning of criteria, including removal of variants with Gnomad allele frequency above 10x10 -06, removal of noncoding variants in low complexity and repetitive regions, those with no functional indications from ANNOVAR annotations, CADD conservative score under 15, and absence in HGMD or COSMIC databases. Highly annotated clinical data, including cytogenetic abnormalities at the latest time point prior to HCT, were obtained from CIBMTR forms. K-means clustering was applied to recurrent mutations and cytogenetic abnormalities to identify clinically relevant genomic subtypes. The optimal cluster number was determined by Gap-status algorithm. Statistics of clinical characteristics were compared among different genomic subgroups by Chi-squared test for categorical variables and Mann-Whitney U test for continuous variables. Overall survival association tests were conducted by Cox multivariate models. Relapse and transplant-related risk were performed by competing risk analysis using Fine-Gray models. Models were adjusted for patient-, disease-, and HCT-related factors.ResultsThe somatic genomic landscape in our MDS cohort was examined for the total count of recurrent mutations at the sample level and gene level. Among 53 recurrently mutated genes in 257 of 494 MDS cases, TP53, TET2, RUNX1, DNMT3A, and ASXL1 were the most frequently mutated genes in our MDS cohort. Based on k-means clustering of the recurrent mutational and cytogenetic data, we detected five clusters that stratified our MDS patient cohort, including one reference cluster with no recurrent somatic mutations or cytogenetic abnormalities. Compared to the reference subgroup, significantly higher cytogenetic scores and IPSS-R scores were observed in genomic clusters with TP53 mutations (cytogenetic score: P=3.42E-07*; IPSS-R score: P= 2.38E-10*) and cytogenetic abnormalities del5q, or tri8p (cytogenetic score: P= 2.38E-10*; IPSS-R score: P=0.09) , or mono7 (cytogenetic score: P=3.29E-13*; IPSS-R score: P=1.38E-05*) (data not shown). Cox multivariate models revealed that genomic clusters with TP53 and del5q mutations (P<0.001*) or tri8p (P=0.02*) mutations have strong associations with post-transplant overall survival outcome (Figure 1A). Furthermore, competing risk analysis confirmed significantly higher risk of relapse in genomic subgroups with TP53 and del5q mutations in the reduced intensity conditioning regimen setting (P=0.01) (Figure 1B), while significantly higher risk of transplant-related mortality was found in the genomic subgroup with tri8p in the myeloablative conditioning regimen setting (P=0.03) (Figure 1C).ConclusionOur study suggests that molecular signatures from MDS patient genomes at HCT may provide an independent prognosis of post-transplant survival. Additionally, our data suggests that the choice of regimen intensity could be informed by knowledge of the individual genomic signature of a given MDS patient. [Display omitted] DisclosuresSaber: Govt. COI: Other.

A unified view of low complexity regions (LCRs) across species

A unified view of low complexity regions (LCRs) across species

The low-complexity domain of the FUS RNA binding protein self-assembles via the mutually exclusive use of two distinct cross-β cores

The low-complexity (LC) domain of the fused in sarcoma (FUS) RNA binding protein self-associates in a manner causing phase separation from an aqueous environment. Incubation of the FUS LC domain under physiologically normal conditions of salt and pH leads to rapid formation of liquid-like droplets that mature into a gel-like state. Both examples of phase separation have enabled reductionist biochemical assays allowing discovery of an N-terminal region of 57 residues that assembles into a labile, cross-β structure. Here we provide evidence of a nonoverlapping, C-terminal region of the FUS LC domain that also forms specific cross-β interactions. We propose that biologic function of the FUS LC domain may operate via the mutually exclusive use of these N- and C-terminal cross-β cores. Neurodegenerative disease-causing mutations in the FUS LC domain are shown to imbalance the two cross-β cores, offering an unanticipated concept of LC domain function and dysfunction.

A vast pool of lineage-specific microproteins encoded by long non-coding RNAs in plants.

Pervasive transcription of eukaryotic genomes results in expression of long non-coding RNAs (lncRNAs) most of which are poorly conserved in evolution and appear to be non-functional. However, some lncRNAs have been shown to perform specific functions, in particular, transcription regulation. Thousands of small open reading frames (smORFs, <100 codons) located on lncRNAs potentially might be translated into peptides or microproteins. We report a comprehensive analysis of the conservation and evolutionary trajectories of lncRNAs-smORFs from the moss Physcomitrium patens across transcriptomes of 479 plant species. Although thousands of smORFs are subject to substantial purifying selection, the majority of the smORFs appear to be evolutionary young and could represent a major pool for functional innovation. Using nanopore RNA sequencing, we show that, on average, the transcriptional level of conserved smORFs is higher than that of non-conserved smORFs. Proteomic analysis confirmed translation of 82 novel species-specific smORFs. Numerous conserved smORFs containing low complexity regions (LCRs) or transmembrane domains were identified, the biological functions of a selected LCR-smORF were demonstrated experimentally. Thus, microproteins encoded by smORFs are a major, functionally diverse component of the plant proteome.