2020单细胞组学国际研讨会 SCO2020

[DawnEve]

时间表

演讲者

[DawnEve]

3.Multiomic Approach to Single Cell Profiling(by Bogdan Budnik)

2020.10.15 Harvard University, FAS, Division of Science.

single cell ProtEomics by Mass Spectrometry

microgial cells co-expressing markers observed in scProteomics.

• IMLR clustering of activated microgilia based on scProteomes analysis.
• Yong and old.

Conformation of sc proteomics finding by alternative technique.

• UMAP of young mice
• tSNE of CyTEK data for

Progress in proteome coverage of original SCoPE MS technique

• Feb 2019 78 cells; May 2019 93cells;

SCREEN(Single Cell pRotEomE aNalysis) MS platform

• Jurkat and U937 cells
• 3x96 plates, detect 4006 proteins.
• Jurkat cell treatment by PMA for 30min vs 4hours:
  • Drug response resolved in time on sc level by SCREEN
  • SCREEN MS platform allow cancer cell type differentiation;
• 磷酸化分析 U937 cell， Doxorubicin 处理后
• 磷酸化分析 U937 cell， activated by Insulin

Towards single cell omics

Can we do metabolomics on proteomics set up?

• 药物处理后的代谢组分析
• BTZ 蛋白酶抑制剂; LBH 组蛋白乙酰化抑制剂; LEN: ...//todo
• Separation of leukemia single cells treated with three drugs based on their metabolomes.
• PCA plot, plate1, column 1: 13,302 compounds; ...
• 显著改变的peaks, control vs drug 处理的组, t-test 改变的峰的数量;

sc omics challenges;

• 便宜 更快, faster
• 蛋白组的深度
• 可重复的样品处理
• 可分析的细胞数量

New frontiers for sc omics, current state for 2019

• SCREEN MS >2900 proteins;
• above 1000 cells per week of ms runs for TMT16plex workflow
• C18柱子做代谢组

Questions. 10:45

Q:
Q: do you use imputation 吗? 做了
Q: Do you do scRNA seq besides Protein omics?
Q: Protein maker is better? or RNA marker?
Protein gives the target for drugs.

[DawnEve]

4.Neutralizing antibodies - The Hope to Cure COVID-19(by Synney Xie)

2020.10.15 10:48

SARS-CoV-2

Prevention
Drugs
RNA virus

Standards for a Cure of COVID19

• 切断传播
• 血浆 太有限了

Enriched memory B cells in a convalescent patient share the identical VDJ sequence but sporadic point mutations

• heavy chain chr14; IGHV2-5, IGHJ3
• light chain chr2;
• 8558 B cells's VDJ, 造了400+重组蛋白，有20个潜在的中和抗体。

[DawnEve]

5.Mapping the dynamic architecture of immune cells in Normality and disease (Nolan Lab)

英语十分流畅、标准

Antibody barcode to cells. Imaging.

TIGIT and PD-L1,

Schematic distinctions and pan-tissue organization

CODEX

• Cutaneous T cell lymphoma (CTCL)
• CTCL CODEX - cellular neighborhood analysis
• CTCL: predictive spatial signature of response

ACE2 gene, COVID-19

• CODEX/MIBI imaging of COVID-19
• MHC1 up in COVID lungs. Present antigen?
• M2 mphs in COVID lungs up;

nultiplexed ion beam imaging

• 5 nm;
• Cisplatin: depleted in nucleus in JQ1_Cisplatin resistance

What to do next?

• What is inevitable?
• AtomScope up-close. 电子分不到待观测物表面，通过一个磁场，达到一个detector上，得到一个3D物体的2D投影。

where we are 2020

• 60-120 marker histochemistry, imaging to the atomic scale
• Neighborhoods &
• Inference of mechanism
• Merging with RNAseq for further mechanism.

Question(28min to 35min)

• Q: 亚群与RNAseq的对应？
• Q:

[DawnEve]

day1 afternoon Session

1.The cell is a bag of RNA(by Steve Quake)

老外，英语标准、流畅。

电生理: Patch-clamp 膜片钳

Unraveling the nature of cellular identity

Voltage and ligand gated ion channel RNA expression matches physiology in slices of mouse ...

Correlation bewteen 电生理和转录组，In slices of mouse hippocampus(海马体)

Drosophila(果蝇) Olfactory(嗅觉器官) System

human Pancreas(胰腺)

Long term memory formation in mice 的转录组学基础

• scRNAseq, tSNE
• seven Activated neuronal subpopulation
• Regulatory motif discovery on differentially expressed genes.

CZ biohub

• the mouse aging cell atlas: tabula muris ... (Nature 2018) (Nature 2020)
  • senescence increases with age;
• Clonality increases with age, B and T cells
• sc data resloves number of cells, versus expression level per cell

狐猴 lemur

Fly cell atlas

Flybase: and all annotation experts.

Cell cycle phase assignments across

• 分化和cycle using RNA velocity and marker-based scoring

Single cell genomics

Qestions(27min to 37)

• Q:

[DawnEve]

day1A: 2.Mapping the human gastrointestinal tract through space and time(by Sarah Teichamann)

sanger;

Gut cell atlas

• Biological insights, disease
• Epithelial cells
• Enteric nervouse system
• Gut-associated lymphoid ...

integrated view of human gut throughout life

• First trimester, second trimester, Pediatric(mucosal biopsy), Adult;
• Building a gut cell census: data from 350,000 cells: UMAP
  • cell group: Epithelial, Endothelial, Mesenchymal, Neuronal, B/Plasma; T/NK; Myeloid; Erythocyte; ...

Global trends in development vs adult

• Questions - Epithelial cells
  • New cell types?
  • BEST4+ Enterocytes.
• Characterising Tuft cells(HPGDS, BMX,FYB1,SH2D7,PLCG2)
  • ProteinAtlas.org

Questions: Neuronal cells

Development of the enteric nervous system

• cell-cycle score: G1, G2M, S

Cell sybtype expression of Hirschsprung's ...

Questions: Gut-associated xx ?(GALT) formation

• CCL21 + CCL19 + activated mLTo: fetal small intestinal & mesent...
• Lymphocyte aggregate at vessels.

Mutation frequency, nascent > adult

Questions:(23min to )

• Q:

[DawnEve]

day1A: 3.Melecular Architecture of the developing brain(by Sten Linarsson)

Wave of differentiation

• age: E6, E10, E14, E18

mouse brain atlas: 18 major classes; 51 subclasses;

• markers: Aif1, Lum, ...

Regionaliztion and patterning of the brain

• Fb, Mb, Hb;

Gliogenesis

Psudolineage tree reconstruction

mousebrain.org

Question(30min to )

[DawnEve]

Fuchou Tang

Lesions ahred mutations, FAP1, different tissues

• cancer epithelial development
• microenvironment

RNA+DNA

• CD45+-, CD3+-
• aneuploidy 非整倍体
• organoid culture 类器官培养

Cancer samples

• 1.single cell squencing - > new Biology
• 2.Organoid culture - > New medicine
• in the process: Gene Editing, drug screening.

[DawnEve]

day2, 1.Chromosome and Genome organization in Pre-implantation development(Peter Fraser)

Weizmann Institute.

Hi-C, sc-Hi-C
老外，英语流畅

Maternal/paternal chromosome volume ratios in 3D

• egg, zygote, 2C, 4C, 8C, 64C
• mother-mother or f-to-f contacts 慢慢变低
• mother-father contacts 慢慢升高，最后达到一致的水平
• Reichman, Science, 2018, zygote 1-cell stage, 3D图

homolog-homolog(同系物) contact?

Whole genome 3D model from a single 4-cell stage cell

• centromeric end
• telemeric end
• 4-CS male balstomere in early -S-phase

What's at the trans-homolog contact sites?

• trans-homolog contacts
• ATAC-seq
• H3K4me3
• H3K27me3*
• H3K9me3
• insulation score
• compartment signal
• miotic hotspots
• ...

genome surveillance?

Suspected explanation for trans-homolog contacts

• B16 genome, (black 16 mouse)
• DBA genome
• Cast genome
• use SNP information(35million?) 分辨小鼠类别

Question(19min to )

• Q: 怎么区分4cell stage的？ Take a pool?
• Q:

[DawnEve]

day2, 2. Comprehensive Brain Map of A Proto-vertebrate(by Mike S. Levine)

Loop vs. Hub

Co-regulation hub

• Shared enhencer: MS2 scyl, ~235kb, PP7 chrb, (Lim et al. Mol Cell 2018)
• Independence of TADs and Tethers: ftz gene?

Lineages of individual neuro..

• stage: iniG, midG, earN, latN, iniT1, earT1, midT2, latT1, latT2, , Lava;
• ddN, Em2

Single Neuron Regulatory Network?

• psudotime heatmap of genes:
• Brain map blueprint

Origins of photoperiodis...

Questions(29min to )

• Q:

[DawnEve]

day2, 3. Epigenomic Atlas of the Mammalian Brain at sc resolution(by Joseph R. Ecker)

甲基化 in 脑

linking Genetic/epigenetic variants to ...

• The cell type(s) impaired by the genetic variants is often unknown.
• Manhattan plot of ... GWAS.

Hypothesis:

• Neuronal types are defined by multipy layers of ...
• multi-modal measurement of single cells
  • Gene expression:
  • chromatin accessibility
  • DNA methylation
  • Chromatin conformation:

Diverse cytosine DNA methylation patterns across human tissues

• Unique non-CG(mCH) DNA methylation in the brain
• CH{CA, CC, CT}

Cytosines Methylation spatial gradients across cortical excitatory neurons

Accurate prediction of cell types and spatial locations using an artificial neural network

• ~24k 100 kb mCH features,
• PCA
• 3000 PCs
• 1000 ANN nodes
• (23min)

Characterizing unique cell types: identifying key transcription factors and motifs

• subtype characteristics in methylome
• 68 Excitatory subtypes
• impact scores(IS), assign pairewise comparison to cell type phylogeny
• motif

Characterizing brain cell types through gene and regulatory...

• Liu, Zhou, et al. (2020) BioRxiv

Gene-Enhencer landscape

• DMR, peaks

Questions(33min to )

• Q:

[DawnEve]

day2, 4.An atalas of Gene Rgulatory elements in adult mouse cerebrum(by Bing Ren)

阿尔兹海默症

Risk variants of late onset of Alzheih... disease

[DawnEve]

Huang Yanyi: CNV of sc

15:34 -

早期建库，时间较长。

• lysis: 1h
• library prep: 打碎、扩增

143 single colon cancer cells(CD45 neg)

• 191 single HEK 293 cells
• 243 single lymphocytes: 2017, a healthy human, 4yr old, male;
• Lymphocyte aneuploid ... FISH

PBMC, Tn5, single cells, Pooling

• 16 patients;
• chrX and chr21;
• age correlation is weak.

3D clear... : Liao et al, PNAS, (2020)

Tn5:

Mineva - A sample-prep method for sequencing COVID-19 clinical ...

• chen, et al, 2020

冷链视频污染可能是COVID-19的传染源

Questions(29min to )

• Q: 怎么区分PCR错误、测序错误，还是PBMC本身的突变？
  • FISH 做验证。还有 ...
• Q: Female double X, express double?

[DawnEve]

Zhang Zemin(16:09 to )

What shapes the diversity of the complex tumor microenvironment(TME)?

Application of single cell sequencing and informatics challenges

• tools

ROGUE(Ratio of global unshifted entropy): a metric for purity

• The largest scRNA-seq database in Omni...

Differences between MSI and MSS CRC T cells

Integration of heterogeneous data(CD8+ T cells)

• Heterogeneity of CD8+ exhausted T cells: TOX, CXCL13, TIGIT, CTLA4, TNFRSF9;
• Two major paths to exhaustion(P1 and P2)
  • P1: through GZMK+ Tem and GZMK+ Tex
  • P2: through ZNF683+Trm;
• The 2 paths are universal among cancer types;
• Clinical implications of T cell-based immune typing;
  • Overall survival

T cell summary

Beyond T cells

Myoloid cells

TAM subtypes identified, with distinct properties.

New myeloid cell types revealed: are they universal?

Pan-cancer analysis of infiltreating myoloid cell ...

Mast cell and DC cell cross-cancer differences.

Distinctive features of tumor-associated macrophages.

肿瘤微环境中免疫细胞的变化

Questions(29min to )

• Q: 不同肿瘤中的免疫细胞比例是否不同？
• Q: 研究T细胞，但是为什么没有研究B或者其他细胞？ 因为是免疫疗法的焦点。现在也研究髓系细胞Myeloid cells.
• Q:

[DawnEve]

Integration of multiple lineage measurements from the same cell reconstructs parallel tumor evolution(by Alexander van Oudenaard) 16:45

老外，英语标准，流畅，较快。 Oncode Institute.

?

ScarTrace: whole-organism clone-tracing using scSeq

为了研究发育，使用内在marker: SNP, SNV

sc multiomics sequencing and analyses of human colorectal cancer: science 2018

Clonal dynamics of colon carcinoma progressing:

Goal: 检测3种 lineage marker in the same single cell during organoid evolution

• CNV:
• Somatic mutation:
• lenti-viral labels at start of experiment:

CNV: in 1641 sc identified 33 distincs copy number variation.

SNP: gSNV, sSNV, 374 cells.

Time: 02,40,60,80,100 代，病毒barcoding 比例的变化 reveals 选择性?

Memorial sloan kettering (MSKCC) database.

Conclusions - organoid evolution

Questions(28min to )

[DawnEve]

the Power of ONE: Immunology in the age of sc genomics(by Ido Amit) 17:25

AMIT LAB

sc for future immunology research

• Quantitative manifold
• activity changes
• genetic perturbations
• dynamics
• evolution
• human pathology
• ...

MARS-seq: massively parallel single cell RNA-seq

• Jaitin et al, science 2014

New tools in the battle against cancer

Giladi et al, Cell 2017

PIC-seq - A novel technology for capturing Immune interactions at sc resolution.

• PIC-seq -> T marker ~ myeloid marker -> TME

immunology beyond cell surface markers.

• Katzen..., cell 2020

INs-sqe: an integrated technology for scRNA-seq and intracellular protein

Questions(34min to )

[DawnEve]

The Cellular Basic for somatosensation and Pain(by Patrik Ernfors) 18:08

a brief history of sensory nerve energies