Molecular Biology in Medicine 医学分子生物学 许正平. The greatest intellectual revolution of the last 40 years may have taken place in biology. Can anyone be considered.

Molecular Biology in Medicine

医学分子生物学

许正平

The greatest intellectual revolution of the last 40 years may have taken place in biology. Can anyone be considered educated today who does not understand a little about molecular biology?

─F. H. Westheimer (Harvard University)

Genetic Information Transfer

遗传信息的传递 Gene Transcription 基因转录 RNA Splicing & Editing RNA 剪切与加工 Protein Synthesis & Processing

蛋白质合成与加工 Regulation of Gene Expression

基因表达的调控 ( 包括 miRNA 、 RNAi)

分子生物学主要内容

分子生物学主要研究技术 分离、纯化（主要是生物大分子） 克隆、表达 PCR （多聚酶链式反应 ） 凝胶电泳：琼脂糖凝胶电泳； SDS －聚丙烯酰胺凝胶电泳 （ SDS-PAGE ）；等电聚焦电泳；双向电泳 印迹技术： Southern blotting; Northern blotting;

Western blotting

微阵列技术： genechip, microarray, protein chip

基因操纵技术： Gene-knockout/knockin

RNA interference (RNAi)

分子生物学主要研究技术 蛋白质相互作用：酵母双杂交、免疫共沉淀（ Co-IP ）、 pull-down 、 FRET 、表面等离子共振技术 (SPR)

蛋白质鉴定：质谱 研究生物大分子三维结构常用的实验手段： X 射线晶体学、核磁共振、电子显微学、原子力显微镜 以及 X 射线小角散射等。

定义：从分子水平研究人体在正常和疾病状态下生命活动及其规律的一门科学重点：人体生物大分子和大分子体系的结构、功能、相互作用及其同疾病发生、发展的关系

医学分子生物学

教材：医学分子生物学（第 3 版）

参考书： Molecular Cell Biology Gene

More Information: Literature Internet

I. Introduction

The Central Dogma

gene expression: The overall process by whichthe information encoded in a gene is converted into an observable phenotype (most commonly production of a protein).

Prokaryotic vs Eukaryotic

I. Introduction

How many genes in the human genome?Gene Expression

I. Introduction

FACT 1: an uniform genome in almost every cell of an organism

FACT 3: the shape and function of each type of cell are different

FACT 2: the proteome in each type of cell is different

I. Introduction

the actions and properties of each cell type are determined by the proteins it contains

FACT 1: an uniform genome in almost every cell of an organism

transcription of different genes largely determines the actions and properties of cells

FACT 3: the shape and function of each type of cell are different

FACT 2: the proteome in each type of cell is different

I. Introduction

the types and amounts of the various proteins in a cell

the concentration of mRNA and the frequency at which the mRNA is translated

which genes are transcribed and their rateof transcription in a particular cell type

TRUTH: the gene is differentially expressed

regulation

same genome in all cells of an organism

regulation

regulation

I. Introduction

Gene Expression Occurs by a Two-Stage Process

Transcription: generates a single-stranded RNA identical in sequence with one of the strands of the duplex DNA Three principal classes of products:

message RNA (mRNA)transfer RNA (tRNA)ribosomal RNA (rRNA)

Principle: complementary base pairing

Translation: converts the nucleotide sequence of an RNA into the sequence of amino acids comprising a protein

each mRNA contains at least one coding region that is related to a protein sequence

II. Transcription

DNA (gene)

RNA polymerase

Regulatory Proteins

Key Players

promoter

A

startpoint terminator

Transcription Unit

template

upstream downstream

enhancer

II. Transcription

Primary transcript is the original unmodified RNA product correspondingto a transcription unit.

Promoter is a region of DNA involved in binding of RNA polymerase to initiate transcription.

RNA polymerases are enzymes that synthesize RNA using a DNA template(formally described as DNA-dependent RNA polymerases).

Terminator is a sequence of DNA, represented at the end of the transcript,that causes RNA polymerase to terminate transcription.

Transcription unit is the distance between sites of initiation and termination by RNA polymerase; may include more than one gene.

Key Terms

Transcription in eukaryotic cells is divided into three classes. Each class is transcribed by a different RNA polymerase:

RNA polymerase I:

RNA polymerase II:

RNA polymerase III:

RNA Polymerase

II. Transcription

Transcription in eukaryotic cells is divided into three classes. Each class is transcribed by a different RNA polymerase:

RNA polymerase I: rRNA; resides in the nucleolus

RNA polymerase II: mRNA, snRNA; locates in the nucleoplasm

RNA polymerase III: tRNA and other small RNAs; nucleoplasm

II. Transcription

RNA Polymerase

The promoters for RNA polymerases I and II are (mostly) upstreamof the startpoint, but some promoters for RNA polymerase III lie downstream of the startpoint.

Each promoter contains characteristic sets of short conserved sequences that are recognized by the appropriate class of factors.

RNA polymerases I and III each recognize a relatively restricted setof promoters, and rely upon a small number of accessory factors. Promoters utilized by RNA polymerase II show more variation in sequence, and are modular in design.

Promoter

II. Transcription

Short sequence elements (cis-acting elements): bind by accessory factors (transcription factors)

The regulatory region might exist in the promoters of certain eukaryotic genes.

Location: usually upstream and in the vicinity of the startpoint.

These sites usually are spread out over a region of >200 bp. common: used constitutivelyspecific: usage is regulated; define a particular class of genes

These sites are organized in different combinations

Cis-acting Element

II. Transcription

Enhancer element is a cis-acting sequence that increases the

utilization of (some) eukaryotic promoters. The components of an enhancer resemble those of the promoter.

Involve in initiation, but far from startpoint. Are targets for tissue-specific or temporal regulation. Function in either orientation and in any location (upstream or

downstream) relative to the promoter.

Enhancer

two characteristics:1. the position of the enhancer need not be

fixed.2. it can function in either orientation.

II. Transcription

promoter enhancer

position fixed variable

action direction one way either orientation

the density of regulatory elements sparse Heavy (closed packed)

redundancy in function no yes

cooperativity between the binding of factors

sequential great

The Difference between Promoter and Enhancer

The distinction between promoters and enhancers is operational, rather than imply a fundamental difference in mechanism

II. Transcription

Most Eukaryotic Genes Are Regulated by Multiple Transcription-Control Elements

(a) Genes of multicellular organisms contain both promoter-proximal elements and enhancersas well as a TATA box or other promoter element. Enhancers may be either upstream or downstream and as far away as 50 kb from the transcription start site. In some cases, promoter-proximal elements occur downstream from the start site as well. (b) Most yeast genes contain only one regulatory region, called an upstream activating sequence (UAS), and a TATA box, which is ≈90 base pairs upstream from the start site.

II. Transcription

Fact: Regulatory elements in eukaryotic DNA often are many kilobases from start sites

Finding Regulatory Element in Eukaryotic DNA

II. Transcription

Transcription Factor

Any protein that is needed for the initiation of transcription, but which is not itself part of RNA polymerase, is defined as a transcription factor.

binds to DNA (trans-acting factor): recognize cis-acting elements

interacts with other protein: recognize RNA pol, or another factor

The common mode of regulation of eukaryotic transcription is positive: a transcription factor is provided under tissue-specific control to activate a promoter or set of promoters that contain a common target sequence. Regulation by specific repression of a target promoter is less common.

II. Transcription

Accessory factors are needed for initiation, principally

responsible for recognizing the promoter.

Interaction with DNA, RNA polymerase, and/or another

factors.

Three groups:

1. General factors

2. Upstream factors

3. Inducible factors

Another name: accessory

II. Transcription

general factors: required for the mechanics of initiating RNA synthesis at all promoters; form a complex surrounding the startpoint with RNA pol, and determine the site of initiation.

basal transcription apparatus (pol + GF)

upstream factors: DNA-binding proteins that recognize specific short consensus elements located upstream of the startpoint. not regulated; ubiquitous; act upon any promoter that contains the appropriate binding site on DNA.

inducible factors: function in the same general way as the upstream factors. have a regulatory role: control transcription patterns in time and space

Accessory Factors

II. Transcription

II. Transcription

Fou

r S

tage

s in

T

ran

scri

pti

on

1. On the genome Which gene(s) to be transcribed? Basic events: Protein binding and/or modification 2. On a specific gene If the gene can be transcribed successfully?

3. On a transcript If the transcript could be correctly spliced? If the transcript could be correctly edited?

Regulation Levels

Key determinant: Cell Signaling!

III. Regulation of transcription

Potential regulation points

Activation of gene structure↓

Initiation of transcription↓

Processing the transcript↓

Termination of transcription↓

Transport to cytoplasm

the overwhelming majority of regulatory events occur at the initiation of transcription

III. Regulation of transcription

5 potential control points:

“Active” Structure

Major Control Point

Alternative Splicing

Regulatory Proteins

the overwhelming majority of regulatory events occur at the initiation of transcription

Key player: regulatory transcription factors

Two questions:1. How does the transcription factor identify its group of target genes?2. How is the activity of the transcription factor itself regulated in response to intrinsic or extrinsic signals?

III. Regulation of transcription

Answer to question 1

The genes share common response element

Structure feature: contain short consensus sequence

Examples:

HSE: heat shock response element; recognized by HSTF

GRE: glucocorticoid response element

SRE: serum response element

MRE: metal response element

III. Regulation of transcription

Regulatory region in MT gene

BLE: basal level element; TRE: TPA response element

General Principle: any one of several different elements, located in either anenhancer or promoter, can independently activate the gene.

III. Regulation of transcription

Answer to question 2

Signal transduction

Key events:

1. Protein synthesis

2. Protein modification

3. Ligand binding

4. Protein cleavage

5. Inhibitor release

6. Mutation

III. Regulation of transcription

The activity of a regulatory transcription factor may be controlled by synthesis of protein, covalent modificationof protein, ligand binding, or binding of inhibitors that sequester the protein or affectits ability to bind to DNA.

Reg

ula

tion

Mod

es o

f T

ran

scri

pti

on

Fac

tor

mutations of the transcriptionfactors give rise to factors thatinappropriately activate, or prevent activation, of

transcription

III. Regulation of transcription

Eukaryotic transcriptional control operates at three levelsduring the stage of initiation

1. changes in chromatin structure directed by activators and repressors

2. modulation of the levels of activators and repressors(gene expression)

3. change the activities of activators and repressors

III. Regulation of transcription

Gene differential expression

IV. RNA Processing

INTRODUCTION

Facts:

1. Genes are interrupted, and mRNAs are uninterrupted

2. The primary transcript has the same organization as the gene

3. mRNA has 5’ cap and 3’ poly(A) tail

4. Heterogeneous nuclear RNAs (hnRNA) exist in the nucleus

5. RNA contains rare bases

Mechanism:

RNA splicing: remove intron

RNA modification: 5’ capping, 3’ polyadenylation, base modification

INTRODUCTION

The initial primary transcript synthesized by RNA polymerase IIundergoes several processing steps before a functional mRNA is produced:

5’ capping 3’ cleavage/polyadenylation RNA splicing

RNA splicing is the process of excising the sequences in RNA that correspond to introns, so that the sequences corresponding to exonsare connected into a continuous mRNA.

IV. RNA Processing

Overview of mRNA Processing in Eukaryotes

The poly(A) tail: ~250 A in mammals, ~150 in insects, ~100 in yeasts. For short primary transcripts with few introns, polyadenylation, cleavage, and splicing usually follows termination. For large genes with multiple introns, introns often are spliced out of the nascentRNA before transcription of the gene is complete.

IV. RNA Processing

Th

e sp

lice

osom

al s

pli

cin

g cy

cle

The splicing snRNPs associate with the pre-mRNA and with eachother in an ordered sequence to form spliceosome

ATP is needed to provide theenergy necessary for rearrangements of the spliceosome structure

IV. RNA Processing

Alternative splicing

Mechanisms:• use of different startpoints or termination sequences• a single primary transcript is spliced in more than one

way, and internal exons are substituted, added, or deleted

Definition: a single gene gives rise to more than one mRNA sequence

Key:what controls the use of such alternative pathways?

IV. RNA Processing

Alternative splicing

Mechanisms:• use of different startpoints or termination sequences• a single primary transcript is spliced in more than one

way, and internal exons are substituted, added, or deleted

Definition: a single gene gives rise to more than one mRNA sequence

Key:what controls the use of such alternative pathways?Protein(s)!

IV. RNA Processing

The Troponin （肌钙蛋白） T (muscle protein) pre-mRNA is alternatively spliced to give rise to

64 different isoforms of the protein

Constitutively spliced exons (exons 1-3, 9-15, and 18)

Mutually exclusive exons (exons 16 and 17)

Alternatively spliced exons (exons 4-8)

Exons 4-8 are spliced in every possible waygiving rise to 32 different possibilities

Exons 16 and 17, which are mutually exclusive,double the possibilities; hence 64 isoforms

IV. RNA Processing

Trans-(intermolecular) splicing

Splicing is usually cis-reaction (intramolecular), but trans-(intermolecular) splicing have been found (very rare). These reactions probably occur by splicesome formation with the appropriate site sequences on each molecule.

trypanosomes and euglenoids: all the mRNAsCaenorhabditis elegans: 10-15% of the mRNAsHuman?

IV. RNA Processing

Initiation of Protein Synthesis

V. Initiation of Protein Synthesis

Critical event:begin protein synthesis at the start codon, thereby setting the stagefor the correct in-frame translation of the entire mRNA.

Main mechanisms:Base pairing between mRNA and rRNABase pairing between mRNA and tRNAMet-tRNAi

Met can only bind at the P site to begin synthesis

Participants: Met-tRNAi

Met

mRNA IFs small subunit large subunit

Protein translation

Two types of methionine tRNA are found in all cells

same aminoacyl-tRNA synthetase (MetRS) charges both tRNAs with methionine

V. Initiation of Protein Synthesis

Eukaryotic initiation of protein synthesis

V. Initiation of Protein Synthesis

Model of protein synthesis on circular polysomes and recycling of ribosomal subunits

PABI and eIF4 (4G and 4E) can interact on mRNA to circularize the molecule

V. Initiation of Protein Synthesis

The nascent polypeptide chain must undergo folding and, in many cases, chemical modification and cleavage to generate the final protein

Folding:Theoretically: any polypeptide chain containing n residues could, in principle, fold into 8n conformations. Fact: adopt a single conformation (native state)

a single, energetically favorable conformation Mechanism: the amino acid sequence provides the information for protein folding

Protein Maturation

Modification: N terminal C terminal Certain sites btw N and C terminus

VI. Protein Processing

Nearly every protein in a cell is chemically altered after its synthesis in a ribosome, thus alter its activity, life span, or cellular location of proteins,

depending on the nature of the alteration.

Two categories:chemical modification involves the linkage of a chemical group to the terminal amino or carboxyl groups or to reactive groups in the side chains of internal residues may be reversibleProcessing involves the removal of peptide segments and generally is irreversible

Protein Alteration

VI. Protein Processing

The internal residues in proteins can be modified by attachment of a variety ofchemical groups to their side chains:phosphorylation (Ser, Thr, Tyr) glycosylation (Asp, Ser, Thr)ubiquitinationothers

Examples of modified internal residuesproduced by hydroxylation, methylation, and carboxylation

Protein Modification

VI. Protein Processing

Protein Cleavage

most common form:

residues are removed from the C- or N-terminus of a polypeptide

by cleavage of the peptide bond in a reaction catalyzed by

proteases.

Proteolytic cleavage is a common mechanism of activation or

inactivation

Proteolysis also generates active peptide hormones

EGF; insulin

VI. Protein Processing

Protein Degradation

Two Pathways

extracellular: digestive proteases

intracellular

lysosomes

cytosolic mechanisms

The ubiquitin-mediated pathway is the best-understood cytosolic pathway.In ubiquitinating enzyme complex, different conjugating enzymes recognize different degradation signals in target proteins.

ubiquitin-conjugating enzyme E1: Arg-X-X-Leu-Gly-X-Ile-Gly-Asxcertain residues at the N-terminus favor rapid ubiquitination

VI. Protein Processing

基因突变、多态性：个体易感性、疾病发生表观遗传修饰：功能改变蛋白质量 / 结构 / 构象 / 功能改变：非正常功能、疾病调控异常：非正常功能、疾病

分子生物学与医学

医学分子生物学的应用

阐明生理 / 病理现象的分子机制 遗传病诊断（基因诊断） 发现疾病的生物标志物，为诊断服务 疾病的生物治疗 药物研发与评价（药物基因组学） 个性化医疗

Tumor Angiogenesisand

The Role of Angiogenin

Part I Tumor Angiogenesis

Angiogenesis

Angiogenesis: the outgrowth of new blood vessels from pre-existing vessels

Normal angiogenesis Tumor angiogenesis

InhibitorsInhibitorsActivatorsActivators

FolkmFolkm

anan

The turn on and off of angiogenesis

The process of angiogenesis

The process of angiogenesis

Tumors can be cured through ‘starving to death’. Avastin (VEGF monoclonal antibody) was approved for combination use with standard chemotherapy for metastatic colon cancer and non-small cell lung cancer in 2004.

Anti-angiogenesis & anti-tumor therapy

Anti-angiogenesis: main target

activation of target cell:ligand-receptor bindingsignalling

cell migrationMMPs

cell adhesion

Ideal target: a specific event in tumor angiogenesis

Anti-angiogenesis: reagent

protein or antibodye.g. avastin

peptide

small molecule

Ideal reagent: no or negligible side-effect

http://www.cancer.gov/cancertopics/factsheet/Therapy/angiogenesis-inhibitors

Anti-angiogenesis: main problem

multi-factor

multi-pathway

side-effect

solution: combined action!

Thalidomide: a scandal

Thalidomide: chiefly sold and prescribed to

pregnant women from 1957 to 1961 in almost

50 countries, as an antiemetic to combat morning

sickness and as an aid to help them sleep.

catastrophic results: severe malformities, including

phocomelia( 海豹肢症 )

Reason: the developing fetus will not form blood

vessels properly and thereby stop the proper

development of fetal limbs and circulatory systems.

Part II The role of angiogenin

Angiogenin (ANG) is one of the major angiogenesis activators Angiogenin (ANG) is one of the major angiogenesis activators

N

C

ANG was originally isolated from the cond

itioned medium of cultured HT-29 human col

on adenocarcinoma cells based solely on its

angiogenic activity.

Mature ANG is a basic, single-chain protei

n containing 123 amino acids with a molecul

ar weight of about 14,400 Da.

ANG gene is present as one copy per hap

loid genome, and localizes on chromosome

14q11.

ANG is a member of ribonuclease superfa

mily.

A brief introduction to angiogenin (ANG)

Angiogenin as a potential drug target

• The expression of angiogenin is up-regulated in various ty

pes of human cancers.

• Many molecules targeting ANG, such as ANG monoclonal

antibodies, have been reported to exert antitumor effects.

• However, they all antagonize angiogenin itself, and may h

ave significant side effect.

• An ideal angiogenin-oriented drug could be made possible

only after fully elucidating the mechanism of action of ang

iogenin and identify a disease-specific process.

Mechanisms of Action of Angiogenin

• Angiogenin induces angiogenesis mainly through:– binding to membrane actin and then inducing basem

ent membrane degradation– binding to a putative 170-kDa protein and subseque

ntly transducting signal into cytoplasm – translocating into the nucleus of target cells directly

and then enhancing rRNA transcription – exerting its ribonucleolytic activity

• Angiogenin can also translocate into the nucleus of cancer cells and induces the corresponding cell proliferation

?

?

?

Research Interest:

Elucidating the mechanism of action of angiogenin

1. Angiogenin-interacting proteins

2. Angiogenin-stimulating miRNAs

Developing anti-angiogenesis and anti-tumor molec

ules

• Since protein interactions are critical in every biologica

l process, interactions between ANG and other proteins

should mediate or modulate a series of biological activi

ties in ANG-induced angiogenesis and tumor cell growt

h.

• The study of angiogenin-interacting proteins will help t

o understand the molecular mechanism of angiogenin-i

nduced angiogenesis and tumorigenesis.

• The study of ANG PPIs may also provide potential targ

ets for anti-angiogenesis therapy.

Why study ANG-interacting proteins?

Yeast Two-hybrid

Co-IP

Approaches employed to study ANG-interacting proteins?

Identify ANG-interacting proteins using yeast two hybrid system

Yeast two-hybrid screening

Interaction confirmation

Mapping the binding sitesExploring the significance of the interaction

Peptide synthesis &Interaction interference

Drug design

Basis of the Two-Hybrid System.(GIBCO BRL® instruction manual)

Principle of yeast two-hybrid

Functional classifications of ANG-binding proteins

ANG-interacting proteins Functional classifications

Laminin

Extracellular matrix proteinsFibronectin

Fibulin1,2,3,4

Latent transforming growth factor-beta binding protein3,4 (LTBP3,4)

Progranulin

Regulatory factors

Follistatin

Sprouty

VE-statin

Chordin

Synovial apoptosis inhibitor 1

Phospholipid Scramblase 1

ANG-interacting proteins Functional classifications

Alpha actinin 2 (ACTN2) Cytoskeleton protein

A Disintegrin And Metalloproteinase protein33 (ADAM33)

Membrane protein

Interferon, alpha-inducible Protein 27 Others

Sarcolemma Associated Protein

FLJ25471, FLJ00221

21 proteins were identified as potential ANG-binding proteins using Y-2-H method.

Functional classifications of ANG-binding proteins

Verification of the potential aniogenin-interacting proteins

reconfirmation in the yeast system

pull-down

in vitro Co-IP

in vivo Co-IP

FRET

a-Actinin-2 (ACTN2), a cytoskeletal protein, binds to angiogenin

ANG ACTN2

- -

- +

+ -

+ +

Identification of ACTN2 as a potential ANG-binding protein in Y-2-H analysis

Detecting the interaction between ACTN2 and ANG using His pull-down

experiment

ACTN

ACTN

His-ACTN2 - + +

WB: ANG

Detecting the interaction between ACTN2 and ANG using Co-Immunoprecipitat

ion experiment

Co-immunoprecipitation experiment

Detecting the interaction between ACTN2 and ANG in vivo using FRET

experiment

Mapping the ANG-binding sites on ACTN2

aa383-632 on ACTN2 was required for ANG binding

Crystal Structure Of The Rod Domain Of Alpha-Actinin

aa383-632 (yellow) on ACTN2 was required for ANG binding

Mapping the ACTN2-binding sites on ANG

aa83-123 on ANG was required for ACTN2 binding

R101 on ANG plays an important role in ANG-ACTN2 interaction

R101

R101 on ANG plays an important role in ANG-ACTN2 interaction

Confirmation of the interaction between Phospholipid Scramblase 1 (PLSCR1) and ANG

PLSCR1 interacts with ANG in yeast PLSCR1 interacts with ANG in vitro

Yeast two-hybrid analysis GST pull-down experiment

Confirmation of the interaction between PLSCR1 and ANG

PLSCR1 interacts with ANG in vivo

PLSCR1 co-localizes with ANG in the nucleolous

PLSCR1 enhances rRNA transcription in HeLa cells

Over-expression of PLSCR1 increases45S rRNA level

Down-regulation of PLSCR1 decreases 45S rRNA level

U6 Ai Ai+ANG

PLSCR1 enhances rRNA transcription through angiogenin

U6: ControlAi: ANGi

ANG expression was down-regulated in ANG RNAigroup and can be recovered with ANG addition

PLSCR1 enhances rRNA transcription only in the presence of ANG

FS interacts with ANG in yeastFS interacts with ANG in yeast

FS interacts with ANG directly in vitroFS interacts with ANG directly in vitro FS interacts with ANG in HeLa cellsFS interacts with ANG in HeLa cells

Identification of the interaction between follistatin (FS) and angiogenin

Follistatin binds to angiogenin through FS1 and FS2 domains

FS inhibits rRNA transcription in HeLa cells

FS localized in the nucleolous of HeLa cells

FS

Nucleolin

Merge

Control FS over-expression

Over-expression of FS decreases45S rRNA level

Follistatin inhibits angiogenin-induced ABE transcription activity

ABE: angiogenin-binding element. It plays an important role in angiogenin-stimulated rRNA transcription.

Principle of Co-Immunoprecipitation

Western Blot or MS

Screen ANG-binding proteins under physiological

circumstance

Flow chart of Co-IP experiment

Cell lysis

ANG+ ANG-

Immunoprecipitation with ANG Abs

MS analysis

Silver staining, looking for differential band

Electrophoresis

Western Blot to verify the binding of ANG and candidate proteins Silver staining of ANG-immunoprecipitates

ANG-interacting proteins Functional classifications

Far upstream element (FUSE) binding protein

Transcription

Y box binding protein 2

Damage-specific DNA binding protein 2

Ubiquitin-like, containing PHD and RING finger domains, 1

SUB1 homolog (S. cerevisiae)

Eukaryotic translation initiation factor 4A, isoform 2

Translation and regulation of protein metabolic process

Tu translation elongation factor, mitochondrial

Heat shock 70kDa protein 9

Functional classifications of ANG-binding proteins

Functional classifications of ANG-binding proteins

Actin, beta

Cytoskeleton, cell motility and junctions

Myosin, heavy chain 9, non-muscle

Alpha actinin 4 (ACTN4)

IQ motif containing GTPase activating protein 1

Spectrin, alpha, erythrocytic 1

Ribonuclease/angiogenin inhibitor 1

Others

Argininosuccinate synthetase 1

Lectin, galactoside-binding, soluble, 12

EF-hand domain family, member D2

T cell receptor beta variable 20-1

Cold inducible RNA binding protein

19 proteins were identified as potential ANG-binding proteins using Co-IP coupled with MS method.

conclusion

• We have identified a bunch of angiogenin-binding partner

s using Y-2-H and Co-IP method.

• Our results showed that PLSCR1 increased ANG-stimulat

ed rRNA transcription while FS acted as a negative regula

tor.

• The significances of the interactions between ANG and ot

her proteins are under study, especially the extracellular

matrix proteins and those related to cell motilities.

PLSCR1FSGRN

Cytoskeleton and cell motility?

?

Thank you for your attention!