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PubMed2024

Protocol to perform integrative analysis of high-dimensional single-cell multimodal data using an interpretable deep learning technique.

The advent of single-cell multi-omics sequencing technology makes it possible for researchers to leverage multiple modalities for individual cells. Here, we present a protocol to perform integrative analysis of high-dimensional single-cell multimodal data using an interpretable deep learning technique called moETM. We describe steps for data preprocessing, multi-omics integration, inclusion of prior pathway knowledge, and cross-omics imputation. As a demonstration, we used the single-cell multi-omics data collected from bone marrow mononuclear cells (GSE194122) as in our original study. For complete details on the use and execution of this protocol, please refer to Zhou et al.1.

Deep Learning↗
PubMed2004

GFP-like proteins as ubiquitous metazoan superfamily: evolution of functional features and structural complexity.

Homologs of the green fluorescent protein (GFP), including the recently described GFP-like domains of certain extracellular matrix proteins in Bilaterian organisms, are remarkably similar at the protein structure level, yet they often perform totally unrelated functions, thereby warranting recognition as a superfamily. Here we describe diverse GFP-like proteins from previously undersampled and completely new sources, including hydromedusae and planktonic Copepoda. In hydromedusae, yellow and nonfluorescent purple proteins were found in addition to greens. Notably, the new yellow protein seems to follow exactly the same structural solution to achieving the yellow color of fluorescence as YFP, an engineered yellow-emitting mutant variant of GFP. The addition of these new sequences made it possible to resolve deep-level phylogenetic relationships within the superfamily. Fluorescence (most likely green) must have already existed in the common ancestor of Cnidaria and Bilateria, and therefore GFP-like proteins may be responsible for fluorescence and/or coloration in virtually any animal. At least 15 color diversification events can be inferred following the maximum parsimony principle in Cnidaria. Origination of red fluorescence and nonfluorescent purple-blue colors on several independent occasions provides a remarkable example of convergent evolution of complex features at the molecular level.

Animals↗
PubMed2001

Water and ion binding around r(UpA)12 and d(TpA)12 oligomers--comparison with RNA and DNA (CpG)12 duplexes.

The structural and dynamic properties of the water and ion first coordination shell of the r(A-U) and d(A-T) base-pairs embedded within the r(UpA)12 and d(TpA)12 duplexes are described on the basis of two 2.4 ns molecular dynamics simulations performed in a neutralizing aqueous environment with 0.25 M added KCl. The results are compared to previous molecular dynamics simulations of the r(CpG)12 and d(CpG)12 structures performed under similar conditions. It can be concluded that: (i) RNA helices are more rigid than DNA helices of identical sequence, as reflected by the fact that RNA duplexes keep their initial A-form shape while DNA duplexes adopt more sequence-specific shapes. (ii) Around these base-pairs, the water molecules occupy 21 to 22 well-defined hydration sites, some of which are partially occupied by potassium ions. (iii) These hydration sites are occupied by an average of 21.9, 21.0, 20.1, and 19.8 solvent molecules (water and ions) around the r(G=C), r(A-U), d(G=C), and d(A-T) pairs, respectively. (iv) From a dynamic point of view, the stability of the hydration shell is the strongest for the r(G=C) pairs and the weakest for the d(A-T) pairs. (v) For RNA, the observed long-lived hydration patterns are essentially non-sequence dependent and involve water bridges located in the deep groove and linking OR atoms of adjacent phosphate groups. Maximum lifetimes are close to 400 ps. (vi) In contrast, for DNA, long-lived hydration patterns are sequence dependent and located in the minor groove. For d(CpG)12, water bridges linking the (G)N3 and (C)O2 with the O4' atoms of adjacent nucleotides with 400 ps maximum lifetimes are characterized while no such bridges are observed for d(TpA)12. (vii) Potassium ions are observed to bind preferentially to deep/major groove atoms at RpY steps, essentially d(GpC), r(GpC), and r(ApU), by forming ion-bridges between electronegative atoms of adjacent base-pairs. On average, about half an ion is observed per base-pair. Positive ion-binding determinants are related to the proximity of two or more electronegative atoms. Negative binding determinants are associated with the electrostatic and steric hindrance due to the proximity of electropositive amino groups and neutral methyl groups. Potassium ions form only transient contacts with phosphate groups.

AT Rich Sequence↗
PubMed2004

Vertical distribution of nitrogen-fixing phylotypes in a meromictic, hypersaline lake.

We investigated the diversity of nitrogenase genes in the alkaline, moderately hypersaline Mono Lake, California to determine (1) whether nitrogen-fixing (diazotrophic) populations were similar to those in other aquatic environments and (2) if there was a pattern of distribution of phylotypes that reflected redox conditions, as well as (3) to identify populations that could be important in N dynamics in this nitrogen-limited lake. Mono Lake has been meromictic for almost a decade and has steep gradients in oxygen and reduced compounds that provide a wide range of aerobic and anaerobic habitats. We amplified a fragment of the nitrogenase gene (nifH) from planktonic DNA samples collected at three depths representing oxygenated surface waters, the oxycline, and anoxic, ammonium-rich deep waters. Forty-three percent of the 90 sequences grouped in nifH Cluster I. The majority of clones (57%) grouped in Cluster III, which contains many known anaerobic bacteria. Cluster I and Cluster III sequences were retrieved at every depth indicating little vertical zonation in sequence types related to the prominent gradients in oxygen and ammonia. One group in Cluster I was found most often at every depth and accounted for 29% of all the clones. These sequences formed a subcluster that contained other environmental clones, but no cultivated representatives. No significant nitrogen fixation was detected by the 15N2 method after 48 h of incubation of surface, oxycline, or deep waters, suggesting that pelagic diazotrophs were contributing little to nitrogen fluxes in the lake. The failure to measure any significant nitrogen fixation, despite the detection of diverse and novel nitrogenase genes throughout the water column, raises interesting questions about the ecological controls on diazotrophy in Mono Lake and the distribution of functional genes in the environment.

Bacteria↗
PubMed2026

Identification of Two Novel Compound Heterozygous ADAMTS17 Variants Associated With Weill-Marchesani Syndrome 4.

PURPOSE: Weill-Marchesani syndrome 4 (WMS4) is frequently underdiagnosed when standard exome sequencing fails to detect noncoding pathogenic variants. We aimed to identify the genetic cause in a patient with suspected WMS4 and to characterize the splicing-altering mechanism of a deep intronic variant in ADAMTS17. MATERIALS AND METHODS: Whole-exome sequencing combined with whole-genome sequencing was conducted in the proband, who presented with ocular and skeletal manifestations of WMS4. A minigene splicing assay was applied to verify the splicing abnormality caused by the deep intronic variant. RESULTS: The patient showed high myopia, brachydactyly, accelerated growth velocity, and advanced bone age. Two novel compound heterozygous variants in ADAMTS17, c.1655G>A and c.450+38C>A, were identified. The deep intronic variant c.450+38C>A was confirmed by minigene assay to disrupt normal pre-messenger RNA splicing by inducing retention of a 35-base pair intronic segment, leading to a frameshift and premature termination (p.G152Lfs*23). CONCLUSIONS: This study broadens the mutational spectrum of ADAMTS17. Whole-genome sequencing combined with functional splicing validation is essential for resolving molecularly undiagnosed cases of WMS4, particularly when deep intronic variants are suspected, and supports clinical genetic testing and genetic counseling of hereditary connective tissue disorders.

ADAMTS17↗
PubMed2025

Deep learning-based cell-specific gene regulatory networks inferred from single-cell multiome data.

Gene regulatory networks (GRNs) provide a global representation of how genetic/genomic information is transferred in living systems and are a key component in understanding genome regulation. Single-cell multiome data provide unprecedented opportunities to reconstruct GRNs at fine-grained resolution. However, the inference of GRNs is hindered by insufficient single omic profiles due to the characteristic high loss rate of single-cell sequencing data. In this study, we developed scMultiomeGRN, a deep learning framework to infer transcription factor (TF) regulatory networks via unique integration of single-cell genomic (single-cell RNA sequencing) and epigenomic (single-cell ATAC sequencing) data. We create scMultiomeGRN to elucidate these networks by conceptualizing TF network graph structures. Specifically, we build modality-specific neighbor aggregators and cross-modal attention modules to learn latent representations of TFs from single-cell multi-omics. We demonstrate that scMultiomeGRN outperforms state-of-the-art models on multiple benchmark datasets involved in diseases and health. Via scMultiomeGRN, we identified Alzheimer's disease-relevant regulatory network of SPI1 and RUNX1 for microglia. In summary, scMultiomeGRN offers a deep learning framework to identify cell type-specific gene regulatory network from single-cell multiome data.

Deep Learning↗
PubMed2026

Human Monocytic Models Reveal Genotype-Dependent Inflammatory Programs in VEXAS Syndrome.

OBJECTIVES: VEXAS syndrome is a severe X-linked autoinflammatory disorder caused by somatic mutations in ubiquitin-like modifier activating enzyme 1 (UBA1), with clinical outcomes that vary by UBA1 genotype. We aimed to elucidate genotype-specific inflammatory programs and identify potential therapeutic targets. METHODS: We conducted longitudinal deep phenotyping, including whole-blood RNA sequencing (RNA-seq) and clinical activity assessment. Peripheral blood samples were analyzed by single-cell RNA-seq. Human monocytic cell lines harboring each major UBA1 mutation (p.Met41Val, p.Met41Thr, or p.Met41Leu) were generated and subjected to transcriptomic and functional analyses. RESULTS: Thirteen patients with VEXAS syndrome contributed a total of 79 RNA-seq samples. Among genes upregulated in VEXAS syndrome, RNASE1 showed the strongest correlation with longitudinal disease activity (r = 0.70, FDR < 0.05) and was upregulated in patients' monocytes. In UBA1-mutant monocytic cell lines, genotype-dependent ubiquitination defects were observed in a graded manner (p.Met41Val > p.Met41Thr > p.Met41Leu), even in the absence of exogenous stimuli. These defects were accompanied by unfolded protein response activation, increased pro-inflammatory cytokine production, progressive cell death, and RNASE1 upregulation, all following the same graded pattern, recapitulating patient genotype-phenotype associations. Transcriptomic analyses demonstrated enrichment of pro-inflammatory, interferon, and necroptosis signatures in more severe genotypes. Notably, inhibition of receptor-interacting protein kinase 3 (RIPK3) markedly attenuated all pathological features, including RNASE1 upregulation. CONCLUSIONS: Our UBA1-mutant monocytic cell-line models, representing three distinct genotypes, recapitulate genotype-dependent inflammatory phenotypes that can be modulated by RIPK3 inhibition, providing a translational platform for mechanistic investigation and precision therapy development in VEXAS syndrome.

Journal Article↗
PubMed2004

OL-protocadherin expression in the visual system of the chicken embryo.

The expression of OL-protocadherin, a homotypically binding cell adhesion molecule, was mapped in the visual system of the chicken embryo at intermediate to late stages of development (11-19 days of incubation). The expression was compared with that of four classic cadherins, described previously. OL-protocadherin is expressed by the isthmooptic nucleus, its retinopetal projection, and possibly its retinal target neurons, the amacrine cells. Ganglion cells begin to express OL-protocadherin at relatively late stages of development. The layers of the optic tectum, the projection neurons in the stratum griseum centrale, and the tectofugal pathways show differential OL-protocadherin immunoreactivity. Several of the diencephalic target nuclei of the tectothalamic projection, such as the principal pretectal nucleus, subpretectal nucleus, and nucleus rotundus, contain distinct subregions or populations of neurons expressing OL-protocadherin. In these centers, the expression pattern of OL-protocadherin differs from that of the four classic cadherins, though it shows partial overlap with them. Other retinorecipient and/or tectorecipient nuclei (ventral geniculate nucleus, lateral dorsolateral nucleus, superficial synencephalic nucleus, pretectal area, and griseum tectale) also show a differential immunoreactivity for OL-protocadherin and other cadherins. Some of these nuclei and the optic tectum display a similar sequence of cadherin expression from superficial to deep layers, in a pattern that may reflect mutual interconnections. This result indicates a partial conservation of cadherin expression across interconnected embryonic divisions, from the mesencephalon to the ventral thalamus. In conclusion, OL-protocadherin is a marker for specific functional gray matter structures and neural circuits in the chicken visual system. J. Comp. Neurol. 470:240-255, 2004.

Animals↗
PubMed2003

Flow-independent magnetic resonance venography of the calf.

PURPOSE: To show that flow-independent venography that exploit the intrinsic MR properties of blood to isolate vessels from surrounding structures can be used for depiction of peripheral veins and for detection of deep venous thrombosis (DVT). MATERIALS AND METHODS: Sequence and parameters were first determined on a theoretical basis. The sequence was then optimized in volunteers (N = 4) for the depiction of the peripheral venous vessels. Qualitative evaluation of the normal venous anatomy was performed in five volunteers. The feasibility of diagnosing DVT of the calf with this method was evaluated with preliminary clinical studies. RESULTS: Excellent depiction of the venous anatomy was achieved in all volunteers with the optimized technique. Very small venous structures, such as superficial, muscular, and perforator veins, were clearly depicted because of the high spatial and contrast resolution capacities of the sequences. In all six patients, DVT findings diagnosed by duplex sonography were also seen on MR venography. CONCLUSION: Venous anatomy mapping and detection of lower-limb DVT appear feasible using flow-independent MR venograms.

Humans↗
PubMed2001

Circularization changes the folding transition state of the src SH3 domain.

Native state topology has been implicated as a major determinant of protein-folding mechanisms. Here, we test experimentally the robustness of the src SH3-domain folding transition state to changes in topology by covalently constraining regions of the protein with disulfide crosslinks and then performing kinetic analysis on point mutations in the context of these modified proteins. Circularization (crosslinking the N and C termini) of the src SH3 domain makes the protein topologically symmetric and causes delocalization of structure in the transition state ensemble suggesting a change in the folding mechanism. In contrast, crosslinking a single structural element (the distal beta-hairpin) which is an essential part of the transition state, results in a protein that folds 30 times faster, but does not change the distribution of structure in the transition state. As the transition states of distantly related SH3 domains were previously found to be very similar, we conclude that the free energy landscape of this protein family contains deep features which are relatively insensitive to sequence variations but can be altered by changes in topology.

Amino Acid Substitution↗
PubMed1999

Molecular phylogenetic relationships of the deep-sea fish genus Coryphaenoides (Gadiformes: Macrouridae) based on mitochondrial DNA.

In order to characterize the phylogenetic relationship and deep-sea adaptation process of the deep-sea fish genus Coryphaenoides, the nucleotide sequences of the mitochondrial (mt) 12 S rRNA and COI gene sequences for seven Coryphaenoides species were analyzed. Our molecular phylogenetic tree shows a new arrangement of seven Coryphaenoides species, which form two distinct groups, abyssal and nonabyssal species, and differs from the results of previous taxonomic studies. Using the mutation rate of mitochondrial genes, the divergence time between abyssal and nonabyssal Coryphaenoides was found to be 3.2-7.6 million years ago. Our study suggests that hydraulic pressure plays an important role in the speciation process in the marine environment.

Adaptation, Physiological↗
PubMed2007

Prokaryotic lifestyles in deep sea habitats.

Gradients of physicochemical factors influence the growth and survival of life in deep-sea environments. Insights into the characteristics of deep marine prokaryotes has greatly benefited from recent progress in whole genome and metagenome sequence analyses. Here we review the current state-of-the-art of deep-sea microbial genomics. Ongoing and future genome-enabled studies will allow for a better understanding of deep-sea evolution, physiology, biochemistry, community structure and nutrient cycling.

Genome↗
PubMed2005

Isoform-specific subcellular localization among 14-3-3 proteins in Arabidopsis seems to be driven by client interactions.

In most higher eukaryotes, the predominantly phosphoprotein-binding 14-3-3 proteins are the products of a multigene family, with many organisms having 10 or more family members. However, current models for 14-3-3/phosphopeptide interactions suggest that there is little specificity among 14-3-3s for diverse phosphopeptide clients. Therefore, the existence of sequence diversity among 14-3-3s within a single organism begs questions regarding the in vivo specificities of the interactions between the various 14-3-3s and their clients. Chief among those questions is, Do the different 14-3-3 isoforms interact with different clients within the same cell? Although the members of the Arabidopsis 14-3-3 family of proteins typically contain highly conserved regions of sequence, they also display distinctive variability with deep evolutionary roots. In the current study, a survey of several Arabidopsis 14-3-3/GFP fusions revealed that 14-3-3s demonstrate distinct and differential patterns of subcellular distribution, by using trichomes and stomate guard cells as in vivo experimental cellular contexts. The effects of client interaction on 14-3-3 localization were further analyzed by disrupting the partnering with peptide and chemical agents. Results indicate that 14-3-3 localization is both isoform specific and highly dependent upon interaction with cellular clients.

14-3-3 Proteins↗
PubMed2025

Rapid Divergence of Visual Systems and Signaling Traits to Contrasting Light Regimes During Early Speciation of African Crater Lake Cichlid Fish.

Sensory adaptation is widely hypothesized to drive ecological speciation, yet empirical evidence from natural populations undergoing early stage divergence remains limited. In Lake Masoko, a young crater lake in East Africa, the haplochromine cichlid Astatotilapia calliptera is undergoing early stage sympatric speciation into shallow-water littoral and deep-water benthic ecotypes that experience contrasting light environments. Here, we integrate retinal transcriptomics, phenotypic analyses, and visual modeling to uncover rapid sensory divergence associated with this ecological transition. We find striking shifts in cone opsin expression, with the benthic ecotype exhibiting a switch from short-wavelength sensitive SWS2B to SWS2A and an overall narrowing of cone sensitivity toward the center of the light spectrum, consistent with changes in deep-water light environment. In contrast, coding sequence variation in opsin genes was limited and no significant differences in allele frequencies were detected across nine polymorphic sites, pointing to expression regulation as the primary axis of early divergence in visual systems. In parallel, we observed divergence in male signaling traits, with benthic males displaying deeper red egg-spots, aligning with predictions from visual modeling of signal efficiency in different light environments. These results demonstrate rapid transcriptomic and phenotypic divergence in associated signaling traits-within &#x223c;1,000 years-supporting a potential role for regulatory evolution in sensory adaptation during early ecological speciation.

Animals↗
PubMed2006

External K+ modulates the activity of the Arabidopsis potassium channel SKOR via an unusual mechanism.

Plant outward-rectifying K+ channels mediate K+ efflux from guard cells during stomatal closure and from root cells into the xylem for root-shoot allocation of potassium (K). Intriguingly, the gating of these channels depends on the extracellular K+ concentration, although the ions carrying the current are derived from inside the cell. This K+ dependence confers a sensitivity to the extracellular K+ concentration ([K+]) that ensures that the channels mediate K+ efflux only, regardless of the [K+] prevailing outside. We investigated the mechanism of K+-dependent gating of the K+ channel SKOR of Arabidopsis by site-directed mutagenesis. Mutations affecting the intrinsic K+ dependence of gating were found to cluster in the pore and within the sixth transmembrane helix (S6), identifying an 'S6 gating domain' deep within the membrane. Mapping the SKOR sequence to the crystal structure of the voltage-dependent K+ channel KvAP from Aeropyrum pernix suggested interaction between the S6 gating domain and the base of the pore helix, a prediction supported by mutations at this site. These results offer a unique insight into the molecular basis for a physiologically important K+-sensory process in plants.

Amino Acid Sequence↗
PubMed2006

Leukoaraiosis is a risk factor for symptomatic intracerebral hemorrhage after thrombolysis for acute stroke.

BACKGROUND AND PURPOSE: The aim of the study was to evaluate whether leukoaraiosis (LA) is a risk factor for symptomatic intracerebral hemorrhage (sICH) in patients treated with thrombolysis for acute stroke. METHODS: In this retrospective, multicenter analysis, we evaluated data from acute anterior circulation stroke patients (n=449; <6 hours after symptom onset) treated with thrombolysis. All patients had received standard magnetic resonance imaging evaluation before thrombolysis, including a high-quality T2-weighted sequence. For the analysis, LA in the deep white matter was dichotomized into absent or mild versus moderate or severe (corresponding to Fazekas scores of 0 to 1 versus 2 to 3). RESULTS: The rate of sICH was significantly more frequent in patients with moderate to severe LA of the deep white matter (n=12 of 114; 10.5%) than in patients without relevant LA (n=13 of 335; 3.8%), corresponding to an odds ratio of 2.9 (95% CI, 1.29 to 6.59; P=0.015). In a logistic-regression analysis (including age, National Institutes of Health Stroke Scale score at presentation, and type of thrombolytic treatment), LA remained a significant independent risk factor (odds ratio, 2.9; P=0.03). CONCLUSIONS: LA of the deep white matter is an independent risk factor for sICH after thrombolytic treatment for acute stroke.

Aged↗
PubMed2024

Shotgun metagenomic analysis of saliva microbiome suggests Mogibacterium as a factor associated with chronic bacterial osteomyelitis.

Osteomyelitis of the jaw is a severe inflammatory disorder that affects bones, and it is categorized into two main types: chronic bacterial and nonbacterial osteomyelitis. Although previous studies have investigated the association between these diseases and the oral microbiome, the specific taxa associated with each disease remain unknown. In this study, we conducted shotgun metagenome sequencing (&#x2265;10 Gb from &#x2265;66,395,670 reads per sample) of bulk DNA extracted from saliva obtained from patients with chronic bacterial osteomyelitis (N = 5) and chronic nonbacterial osteomyelitis (N = 10). We then compared the taxonomic composition of the metagenome in terms of both taxonomic and sequence abundances with that of healthy controls (N = 5). Taxonomic profiling revealed a statistically significant increase in both the taxonomic and sequence abundance of Mogibacterium in cases of chronic bacterial osteomyelitis; however, such enrichment was not observed in chronic nonbacterial osteomyelitis. We also compared a previously reported core saliva microbiome (59 genera) with our data and found that out of the 74 genera detected in this study, 47 (including Mogibacterium) were not included in the previous meta-analysis. Additionally, we analyzed a core-genome tree of Mogibacterium from chronic bacterial osteomyelitis and healthy control samples along with a reference complete genome and found that Mogibacterium from both groups was indistinguishable at the core-genome and pan-genome levels. Although limited by the small sample size, our study provides novel evidence of a significant increase in Mogibacterium abundance in the chronic bacterial osteomyelitis group. Moreover, our study presents a comparative analysis of the taxonomic and sequence abundances of all genera detected using deep salivary shotgun metagenome data. The distinct enrichment of Mogibacterium suggests its potential as a marker to distinguish between patients with chronic nonbacterial osteomyelitis and chronic bacterial osteomyelitis, particularly at the early stages when differences are unclear.

Humans↗
PubMed2005

Phylogenetic analysis of Archaea in the deep-sea sediments of west Pacific Warm Pool.

Archaea are known to play important roles in carbon cycling in marine sediments. The main compositions of archaeal community in five deep-sea sediment samples collected from west Pacific Warm Pool area (WP-0, WP-1, WP-2, WP-3, WP-4), and in five sediment layers (1 cm-, 3 cm-, 6 cm-, 10 cm-, 12 cm- layer) of the 12 cm sediment core of WP-0 were checked and compared by denaturing gradient gel electrophoresis and 16 S rRNA gene sequencing. It was revealed that all the deep-sea sediment samples checked contained members of non-thermophilic marine group I crenarchaeota as the predominant archaeal group. To further detect groups of archaea possibly relating with C1 metabolism, PCR amplification was carried out using primers targeting methane-oxidizing archaea. Although no methane-oxidizing archaea was detected, a group of novel archaea (named as WPA) was instead identified from all these five WP samples by clone analysis. They could be placed in the euryarchaeota kingdom, separated into two distinct groups, the main group was peripherally related with methanogens, the other group related with Thermoplasma. The vertical distributions of WPA, archaea and bacteria along the WP-0 sediment column were determined by quantitative-PCR. It was found that bacteria dominated at all depths, the numbers of bacteria were 10-10(4) times more than those of archaea. The proportion of archaea versus bacteria had a depth related increasing tendency, it was lowest at the first layer (0.01%), reached highest at the 12 cm- layer (10%). WPA only constituted a small proportion of the archaeal community (0.05% to 5%) of west Pacific Warm Pool sediment.

Archaea↗