Biology subjects

Yu, Y.

Publications and source records attributed to Yu, Y..

At least 271 records · Page 15Linked to original sources

bioRxiv2024

Integrated analysis and systematic characterization of the regulatory network for human germline development

Primordial germ cells (PGCs) are the precursors of germline that are specified at embryonic stage. Recent studies reveal that humans employ different mechanisms for PGC specification compared with model organisms such as mice. Moreover, the specific regulatory machinery is still largely unexplored, mainly due to the inaccessible nature of this complex biological process. Here, we collect and integrate multi-omics data, including 581 RNA-seq, 54 ATAC-seq, 45 ChIP-seq, and 69 single-cell RNA-seq samples from different stages of human PGC development to recapitulate the precisely controlled and stepwise process, presenting an atlas in the human PGC database (hPGCdb). With these uniformly processed data and integrated analyses, we characterize the potential key transcription factors and regulatory networks governing human germ cell fate. We validate the important roles of some of the key factors in germ cell development by CRISPRi knockdown. We also identify the soma-germline interaction network and discover the involvement of SDC2 and LAMA4 for PGC development, as well as soma-derived NOTCH2 signaling for germ cell differentiation. Taken together, we have built a database for human PGCs and demonstrate that hPGCdb enables the identification of the missing pieces of mechanisms governing germline development, including both intrinsic and extrinsic regulatory programs.

developmental biology↗

bioRxiv2024

De novo multi-mechanism antimicrobial peptide design via multimodal deep learning

Artificial intelligence (AI)-driven discovery of antimicrobial peptides (AMPs) is yet to fully utilise their three-dimensional (3D) structural characteristics, microbial specie-specific antimicrobial activities and mechanisms. Here, we constructed a QLAPD database comprising the sequence, structures and antimicrobial properties of 12,914 AMPs. QLAPD underlies a multimodal, multitask, multilabel, and conditionally controlled AMP discovery (M3-CAD) pipeline, which is proposed for the de novo design of multi-mechanism AMPs to combat multidrug-resistant organisms (MDROs). This pipeline integrates the generation, regression, and classification modules, using a innovative 3D voxel coloring method to capture the nuanced physicochemical context of amino acids, significantly enhancing structural characterizations. QL-AMP-1, discovered by M3-CAD, which possesses four antimicrobial mechanisms, exhibited low toxicity and significant activity against MDROs. The skin wound infection model demonstrates its considerable antimicrobial effects and negligible toxicity. Altogether, integrating 3D features, specie-specific antimicrobial activities and mechanisms enhanced AI-driven AMP discovery, making the M3-CAD pipeline a viable tool for de novo AMP design.

bioengineering↗

bioRxiv2023

SECE: accurate identification of spatial domain by incorporating global spatial proximity and local expression proximity

MotivationAccurate identification of spatial domains is essential for analyzing spatial transcriptomics data to elucidate tissue microenvironments and biological functions. Existing methods utilize either local or global spatial relationships between spots to aid domain segmentation. A method that can concurrently capture both local and global spatial information may improve identification of spatial domains. ResultsIn this article, we propose SECE, a deep learning-based method that captures both local and global relationships among spots and aggregates their information using expression similarity and spatial similarity. We benchmarked SECE against eight state-of-the-art methods on six real spatial transcriptomics datasets spanning four different platforms. SECE consistently outperformed other methods in spatial domain identification accuracy. Moreover, SECE produced spatial embeddings that exhibited clearer patterns in low-dimensional visualizations and facilitated more accurate trajectory inference. Availability and implementationSECE is implemented and provided as a pip installable Python package which is available on GitHub https://github.com/xie-lab/SECE.

bioinformatics↗

bioRxiv2023

Visual information is broadcast among cortical areas in discrete channels

Withdrawal StatementThe authors have withdrawn this manuscript due to a duplicate posting of manuscript number BIORXIV/2018/469114. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author. The correct preprint can be found at doi: 10.1101/469114

neuroscience↗

bioRxiv2023

PEARL-catalyzed peptide bond formation after chain reversal during the biosynthesis of non-ribosomal peptides

A subset of nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs) are encoded in their biosynthetic gene clusters (BGCs) with enzymes annotated as lantibiotic dehydratases. The functions of these putative lantibiotic dehydratases remain unknown. Here, we characterize an NRPS-PKS BGC with a putative lantibiotic dehydratase from the bacterium Stackebrandtia nassauensis (sna). Heterologous expression revealed several metabolites produced by the BGC, and the omission of selected biosynthetic enzymes revealed the biosynthetic sequence towards these compounds. The putative lantibiotic dehydratase catalyzes peptide bond formation that extends the peptide scaffold opposite to the NRPS and PKS biosynthetic direction. The condensation domain of the NRPS catalyzes the formation of a ureido group, and bioinformatics analysis revealed distinct active site residues of ureido-generating condensation (UreaC) domains. This work demonstrates that the annotated lantibiotic dehydratase serves as a separate amide bond-forming machinery in addition to the NRPS, and that the lantibiotic dehydratase enzyme family possesses diverse catalytic activities in the biosynthesis of both ribosomal and non-ribosomal natural products.

biochemistry↗

bioRxiv2023

Inhibition of O-GlcNAc transferase activates type I interferon-dependent antitumor immunity by bridging cGAS-STING pathway

The O-GlcNAc transferase (OGT) is an essential enzyme that mediates protein O-GlcNAcylation, a unique form of posttranslational modification of many nuclear and cytosolic proteins. Recent studies observed increased OGT and O-GlcNAcylation levels in a broad range of human cancer tissues compared to adjacent normal tissues, indicating a universal effect of OGT in promoting tumorigenesis. Here, we show that OGT is essential for tumor growth in immunocompetent hosts by repressing the cyclic GMP-AMP synthase (cGAS)-dependent DNA sensing pathway. We found that deletion of OGT (Ogt-/-) caused a marked reduction in tumor growth in both syngeneic tumor models and a genetic colorectal cancer (CRC) model induced by mutation of the Apc gene (Apcmin). Pharmacological inhibition or genetic deletion of OGT induced a robust genomic instability (GIN), leading to cGAS-dependent production of the type I interferon (IFN-I) and IFN-stimulated genes (ISGs). As a result, deletion of Cgas or Sting from Ogt-/- cancer cells restored tumor growth, and this correlated with impaired CD8+ T cell-mediated antitumor immunity. Mechanistically, we found that OGT-dependent cleavage of host cell factor C1 (HCF-1) is required for the avoidance of GIN and IFN-I production in tumors. In summary, our results identify OGT-mediated genomic stability and activate cGAS-STING pathway as an important tumor cell-intrinsic mechanism to repress antitumor immunity.

cancer biology↗

bioRxiv2023

Electron-bifurcation and fluoride efflux systems in Acetobacterium spp. drive defluorination of perfluorinated unsaturated carboxylic acids

Enzymatic cleavage of C-F bonds in per- and polyfluoroalkyl substances (PFAS) is largely unknown but avidly sought to promote systems biology for PFAS bioremediation. Here, we report the reductive defluorination of , {beta}-unsaturated per- and polyfluorocarboxylic acids by Acetobacterium spp. Two critical molecular features in Acetobacterium species enabling reductive defluorination are (i) a functional fluoride efflux transporter (CrcB) and (ii) an electron-bifurcating caffeate reduction pathway (CarABCDE). The fluoride transporter was required for detoxification of released fluoride. Car enzymes were implicated in defluorination by the following evidence: (i) only Acetobacterium spp. with car genes catalyzed defluorination; (ii) caffeate and PFAS competed in vivo; (iii) models from the X-ray structure of the electron-bifurcating reductase (CarC) positioned the PFAS substrate optimally for reductive defluorination; (iv) products identified by 19F-NMR and high-resolution mass spectrometry were consistent with the model. Defluorination biomarkers identified here were found in wastewater treatment plant metagenomes on six continents.

microbiology↗

bioRxiv2023

Genomic Revisitation and Reclassification of the Genus Providencia

Members of Providencia, although typically opportunistic, can cause severe infections in immunocompromised hosts. Recent advances in genome sequencing provide an opportunity for more precise study of this genus. In this study, we first identified and characterized a novel species named Providencia zhijiangensis sp. nov. It has [≤]88.23% average nucleotide identity (ANI) and [≤]31.8% in silico DNA-DNA hybridization (dDDH) values with all known Providencia species, which fall significantly below the species-defining thresholds. Interestingly, we found that Providencia stuartii and Providencia thailandensis actually fall under the same species, evidenced by an ANI of 98.59% and a dDDH value of 90.4%. By fusing ANI with phylogeny, we have reclassified 545 genomes within this genus into 20 species, including seven unnamed taxa (provisionally titled Taxon1-7), which can be further subdivided into 23 lineages. Pangenomic analysis identified 1,550 genus-core genes in Providencia, with coenzymes being the predominant category at 10.56%, suggesting significant intermediate metabolism activity. Resistance analysis revealed that most lineages of the genus (82.61%, 19/23) carry a high number of antibiotic resistance genes (ARGs) and display diverse resistance profiles. Notably, the majority of ARGs are located on plasmids, underscoring the significant role of plasmids in the resistance evolution within this genus. Three species or lineages (P. stuartii, Taxon 3, and Providencia hangzhouensis L12) that possess the highest number of carbapenem resistance genes suggest their potential influence on clinical treatment. These findings underscore the need for continued surveillance and study of this genus, particularly due to their role in harboring antibiotic resistance genes.

microbiology↗

bioRxiv2023

Yeast EndoG prevents genome instability by degrading cytoplasmic DNA

In metazoans release of mitochondrial DNA or retrotransposon cDNA to cytoplasm can cause sterile inflammation and disease. Cytoplasmic nucleases degrade these DNA species to limit inflammation. It remains unknown whether degradation these DNA also prevents nuclear genome instability. To address this question, we decided to identify the nuclease regulating transfer of these cytoplasmic DNA species to the nucleus. We used an amplicon sequencing-based method in yeast enabling analysis of millions of DSB repair products. Nuclear mtDNA (NUMTs) and retrotransposon cDNA insertions increase dramatically in nondividing stationary phase cells. Yeast EndoG (Nuc1) nuclease limits insertions of cDNA and transfer of very long mtDNA (>10 kb) that forms unstable circles or rarely insert in the genome, but it promotes formation of short NUMTs ([~]45-200 bp). Nuc1 also regulates transfer of cytoplasmic DNA to nucleus in aging or during meiosis. We propose that Nuc1 preserves genome stability by degrading retrotransposon cDNA and long mtDNA, while short NUMTs can originate from incompletely degraded mtDNA. This work suggests that nucleases eliminating cytoplasmic DNA play a role in preserving genome stability.

genetics↗

bioRxiv2023

Toward Best Practice in Identifying Subtle Differential Expression with RNA-seq: A Real-World Multi-Center Benchmarking Study Using Quartet and MAQC Reference Materials

Translating RNA-seq into clinical diagnostics requires ensuring the reliability of detecting clinically relevant subtle differential expressions, such as those between different disease subtypes or stages. Moreover, cross-laboratory reproducibility and consistency under diverse experimental and bioinformatics workflows urgently need to be addressed. As part of the Quartet project, we presented a comprehensive RNA-seq benchmarking study utilizing Quartet and MAQC RNA reference samples spiked with ERCC controls in 45 independent laboratories, each employing their in-house RNA-seq workflows. We assessed the data quality, accuracy and reproducibility of gene expression and differential gene expression and compared over 40 experimental processes and 140 combined differential analysis pipelines based on multiple ground truths. Here we show that real-world RNA-seq exhibited greater inter-laboratory variations when detecting subtle differential expressions between Quartet samples. Experimental factors including mRNA enrichment methods and strandedness, and each bioinformatics step, particularly normalization, emerged as primary sources of variations in gene expression and have a more pronounced impact on the subtle differential expression measurement. We underscored the pivotal role of experimental execution over the choice of experimental protocols, the importance of strategies for filtering low-expression genes, and optimal gene annotation and analysis tools. In summary, this study provided best practice recommendations for the development, optimization, and quality control of RNA-seq for clinical diagnostic purposes.

genomics↗

bioRxiv2023

Lithocholic acid targets TULP3 to activate sirtuins and AMPK to retard ageing

Lithocholic acid (LCA), accumulated in the body during calorie restriction (CR), can confer administered metazoans with the ability to activate AMP-activated protein kinase (AMPK) and retard ageing. However, how LCA is signalled to activate AMPK and elicit the biological effects is unclear. Here, we show that LCA can enhance sirtuins (SIRTs) to deacetylate and subsequently inhibit vacuolar H+-ATPase (v-ATPase), thereby triggering AMPK activation via the lysosomal glucose-sensing pathway. Through proteomic analysis of SIRT1-coimmunoprecipitated proteins, we identify and validate that TUB like protein 3 (TULP3) is a constitutive component of SIRTs. Surprisingly, we found that TULP3 is an LCA receptor, and that the LCA-bound TULP3 activates SIRTs. The activated SIRTs in turn deacetylate the V1E1 subunit of v-ATPase on K52, K99 and K191 residues. Muscle-specific expression of the 3KR mutant of V1E1, mimicking the deacetylated state, dominantly activates AMPK and rejuvenates muscles in aged mice. Moreover, LCA once administered also activates AMPK and extends lifespan and healthspan in nematodes and flies, depending on the TULP3 homologues tub-1 and ktub, respectively. Our study thus elucidates that LCA triggers the TULP3-sirtuin-v-ATPase- AMPK route to manifest benefits of calorie restriction.

physiology↗

bioRxiv2023

Large-Scale Information Retrieval and Correction of Noisy Pharmacogenomic Datasets through Residual Thresholded Deep Matrix Factorization

Pharmacogenomics studies are attracting an increasing amount of interest from researchers in precision medicine. The advances in high-throughput experiments and multiplexed approaches allow the large-scale quantification of drug sensitivities in molecularly characterized cancer cell lines (CCLs), resulting in a number of open drug sensitivity datasets for drug biomarker discovery. However, a significant inconsistency in drug sensitivity values among these datasets has been noted. Such inconsistency indicates the presence of substantial noise, subsequently hindering downstream analyses. To address the noise in drug sensitivity data, we introduce a robust and scalable deep learning framework, Residual Thresholded Deep Matrix Factorization (RT-DMF). This method takes a single drug sensitivity data matrix as its sole input and outputs a corrected and imputed matrix. Deep Matrix Factorization (DMF) excels at uncovering subtle patterns, due to its minimal reliance on data structure assumptions. This attribute significantly boosts DMFs ability to identify complex hidden patterns among nuisance effects in the data, thereby facilitating the detection of signals that are therapeutically relevant. Furthermore, RT-DMF incorporates an iterative residual thresholding (RT) procedure, which plays a crucial role in retaining signals more likely to hold therapeutic importance. Validation using simulated datasets and real pharmacogenomics datasets demonstrates the effectiveness of our approach in correcting noise and imputing missing data in drug sensitivity datasets (open source package available at https://github.com/tomwhoooo/rtdmf).

bioinformatics↗

bioRxiv2023

Identification and validation of aging-related genes in heart failure based on multiple machine learning algorithms

BackgroundIn the face of continued growth in the elderly population, the need to understand and combat age-related cardiac decline becomes even more urgent, requiring us to uncover new pathological and cardioprotective pathways. MethodsWe obtained the aging-related genes of heart failure through WGCNA and CellAge database. We elucidated the biological functions and signaling pathways involved in heart failure and aging through GO and KEGG enrichment analysis. We used three machine learning algorithms: LASSO, RF and SVM-RFE to further screen the aging-related genes of heart failure, and fitted and verified them through a variety of machine learning algorithms. Finally, we searched for drugs to treat age-related heart failure through the DSigDB database. ResultsWe obtained 57 up-regulated and 195 down-regulated aging-related genes in heart failure through WGCNA and CellAge databases. GO and KEGG enrichment analysis showed that aging-related genes are mainly involved in mechanisms such as Cellular senescence and Cell cycle. We further screened aging-related genes through machine learning and obtained 14 key genes. We verified the results on the test set and 2 external validation sets using 15 machine learning algorithm models and 207 combinations, and the highest accuracy was 0.911. Through screening of the DSigDB database, we believe that rimonabant and lovastatin have the potential to delay aging and protect the heart. ConclusionsWe identified aging signature genes and potential therapeutic drugs for heart failure through bioinformatics and multiple machine learning algorithms, providing new ideas for studying the mechanism and treatment of age-related cardiac decline.

bioinformatics↗

bioRxiv2023

FITM2 deficiency results in ER lipid accumulation, ER stress, reduced apolipoprotein B lipidation, and VLDL triglyceride secretion in vitro and in mouse liver

Structured AbstractO_ST_ABSObjectivesC_ST_ABSTriglyceride (TG) association with apolipoprotein B100 (apoB100) serves to form very low density lipoproteins (VLDL) in the liver. The repertoire of factors that facilitate this association is incompletely defined. FITM2, an integral endoplasmic reticulum (ER) protein, was originally discovered as a factor participating in cytoplasmic lipid droplets (LDs) in tissues that do not form VLDL. We hypothesized that in the liver, in addition to promoting cytosolic LD formation, FITM2 would also transfer TG from its site of synthesis in the ER membrane to nascent VLDL particles within the ER lumen. MethodsExperiments were conducted using a rat hepatic cell line (McArdle-RH7777, or McA cells), an established model of mammalian lipoprotein metabolism, and mice. FITM2 expression was reduced using siRNA in cells and by liver specific cre-recombinase mediated deletion of the Fitm2 gene in mice. Effects of FITM2 deficiency on VLDL assembly and secretion in vitro and in vivo were measured by multiple methods, including density gradient ultracentrifugation, chromatography, mass spectrometry, simulated Raman spectroscopy (SRS) microscopy, sub-cellular fractionation, immunoprecipitation, immunofluorescence, and electron microscopy. Main findings1) FITM2-deficient hepatic cells in vitro and in vivo secrete TG-depleted VLDL particles, but the number of particles is unchanged compared to controls; 2) FITM2 deficiency in mice on a high fat diet (HFD) results in decreased plasma TG levels. The number of apoB100-containing lipoproteins remains similar, but shift from VLDL to LDL density; 3) Both in vitro and in vivo, when TG synthesis is stimulated and FITM2 is deficient, TG accumulates in the ER, and despite its availability this pool is unable to fully lipidate apoB100 particles; 4) FITM2 deficiency disrupts ER morphology and results in ER stress. Principal conclusionsThe results suggest that FITM2 contributes to VLDL lipidation, especially when newly synthesized hepatic TG is in abundance. In addition to its fundamental importance in VLDL assembly, the results also suggest that under dysmetabolic conditions, FITM2 may be a limiting factor that ultimately contributes to non-alcoholic fatty liver disease (NAFLD) and steatohepatitis (NASH).

biochemistry↗

bioRxiv2023

Postural Stability During Illusory Self-Motion - Interactions of Vision and Touch

The role of vision in stabilizing balance has been studied exhaustively. Other studies have shown that non-supportive light touch of the fingertip with a surface also can significantly stabilize postural balance. We have studied how vision and cutaneous information jointly affect balance. We used a head-mounted display to simulate a virtual room that rotated about a vertical axis centered with the standing subjects z-axis. Subjects viewing the displays rotational displacement soon experienced self-motion and displacement. We assessed how the moving visual input destabilized posture and how it interacted with touch cues that stabilized posture. A novel result is how balance is influenced by the onset of visual motion and the illusion of self-rotation. We discovered that motion perceptions are coupled with stochastic aspects of balance. Changes in the perception of types of motion - none, environment-, and self-rotation - distinctively influence metrics that encode for the stochasticity of balance and do not influence those that filter the stochasticity out. We reconfirmed the significant effects of touch in stabilizing balance and discovered how it interacts with the visual perception of motion. We also found lingering effects of past motion perception, which keep influencing the stochasticity of balance even when visual motion is long stopped. Our findings provide insights into multisensory interaction effects in postural balance and suggest novel future research directions.

neuroscience↗

bioRxiv2023

Transfer of disulfide bond formation modules via yeast artificial chromosomes promotes the expression of heterologous proteins in Kluyveromyces marxianus

Kluyveromyces marxianus is a food-safe yeast with great potential for producing heterologous proteins. Improving the yield in K. marxianus remains a challenge, while incorporating large-scale functional modules poses a technical obstacle in engineering. To address these issues, linear and circular yeast artificial chromosomes of K. marxianus (KmYACs) were constructed and loaded with disulfide bond formation modules from Pichia pastoris or K. marxianus. These modules contained up to 7 genes with a maximum size of 15 kb. KmYACs carried telomeres either from K. marxianus or Tetrahymena. KmYACs were transferred successfully into K. marxianus and stably propagated without affecting the normal growth of the host, regardless of the type of telomeres and configurations of KmYACs. KmYACs increased the overall expressions of disulfide bond formation genes and significantly enhanced the yield of various heterologous proteins. In high-density fermentation, the use of KmYACs resulted in a glucoamylase yield of 16.8 g/L, the highest reported level to date in K. marxianus. Transcriptomic and metabolomic analysis of cells containing KmYACs suggested increased FAD biosynthesis, enhanced flux entering the TCA cycle and a preferred demand for lysine and arginine as features of cells overexpressing heterologous proteins. Consistently, supplementing lysine or arginine further improved the yield. Therefore, KmYAC provides a powerful platform for manipulating large modules with enormous potential for industrial applications and fundamental research. Transferring the disulfide bond formation module via YACs proves to be an efficient strategy for improving the yield of heterologous proteins, and this strategy may be applied to optimize other microbial cell factories. Impact StatementIn this study, yeast artificial chromosomes of K. marxianus (KmYACs) were constructed and successfully incorporating modules for large-scale disulfide bond formation. KmYACs were stably propagated in K. marxianus without compromising the normal growth of the host, irrespective of the selection of telomeres (either Tetrahymena or K. marxianus) and configuration (either linear or circular). KmYACs notably enhanced the expressions of various heterologous proteins, with further yield improvement by supplementing lysine or arginine in the medium. Our findings affirm KmYAC as a robust and versatile platform for transferring large-scale function modules, demonstrating immense potential for both industrial applications and fundamental research.

synthetic biology↗

bioRxiv2023

The Effects of Different Fatigue Types on Action Anticipation and Physical Performance in High-level Volleyball Players

This study examined the effects of different fatigue types on action anticipation and physical performance in high-level volleyball players. Twenty-four participants underwent four counterbalanced conditions: 60-min of cycling at 60% peak power output, 60-min Stroop task, 60-min cycling at 60% peak power output with Stroop task and 60-min neutral documentary to induce physical fatigue (PF), mental fatigue (MF), dual fatigue (DF) and control group (CG), respectively. Action anticipation (anticipation test and visual search test) and physical performance (countermovement jump, T-test, and spike test) were conducted at baseline, immediately after (Post1), and 40-min after fatigue intervention (Post2). DF and PF led to significantly lower jump height, T-test completion time, and spiking speed at Post1 and Post2 compared with CG and MF. Compared with PF, DF led to a significant decline in jumping, agility, and spiking performance at Post1 and decreased jumping performance at Post2. MF significantly decreased reaction time in the anticipation test at Post1 compared with CG. Reaction times in the visual search test were significantly lower with DF and MF at Post2 compared with CG. PF led to a greater decline in physical performance and this was exacerbated in DF, yet anticipation performance remained lesser impact.

physiology↗

bioRxiv2023

Fast evolution of SARS-CoV-2 BA.2.86 to JN.1 under heavy immune pressure

While the BA.2.86 variant demonstrated significant antigenic drift and enhanced ACE2 binding affinity, its ability to evade humoral immunity was relatively moderate compared to dominant strains like EG.5 and HK.3. However, the emergence of a new subvariant, JN.1 (BA.2.86.1.1), which possesses an additional spike mutation, L455S, compared to BA.2.86, showed a markedly increased prevalence in Europe and North America, especially in France. Here, we found that L455S of JN.1 significantly enhances immune evasion capabilities at the expense of reduced ACE2 binding affinity. This mutation enables JN.1 to effectively evade Class 1 neutralizing antibodies, offsetting BA.2.86s susceptibility and thus allowing it to outcompete both its precursor BA.2.86 and the prevailing variants HV.1 (XBB.1.5+L452R+F456L) and JD.1.1 (XBB.1.5+L455F+F456L+A475V) in terms of humoral immune evasion. The rapid evolution from BA.2.86 to JN.1, similar to the earlier transition from BA.2.75 to CH.1.1, highlights the importance of closely monitoring strains with high ACE2 binding affinity and distinct antigenicity, despite their temporarily unremarkable immune evasion capabilities. Such strains could survive and transmit at low levels, since their large antigenic distance to dominant strains allow them to target distinct populations and accumulate immune-evasive mutations rapidly, often at the cost of receptor binding affinity.

immunology↗