RESULT: The gene is the HUMAN c-H-ras gene
Compare sequence to NORMAL gene
RESULT: RAS “Activation” is due to a SINGLE point mutation (gly val) at codon 12
Mouse
Drosophila
Human
+
+
+
+
+
Thus: a proto-oncogene is the NORMAL progenitor gene of a viral oncogene
1980, 1st oncogene identified
1989 Nobel Prize for Bishop and Varmus
Oncogene: an altered gene whose product can act in a dominant fashion to help
make a cell cancerous. Typically, an oncogene is a mutant form of a normal gene (proto-oncogene) involved in the control of cell growth or division.
C Isolation of Retroviral Transforming Genes
RSV (gag, pol, env, src)
RSV (td: gag, pol, env)
R.T.(1975 Nobel)
cDNA
genomic RNA
gag, pol env src
Denature and Hybridize 1975,1977
一、癌基因 Oncogene
1.癌基因的发现
Virus and Cancer
A Non-Transforming (Avian Leukemia Viruses, ALV)
Ellerman and Bang, 1908 1, Induce leukemia after long latency periods 2, Do not “transform” tissue culture cells
gag, pol, env
unhybridized sequences
hybridized sequences
src
gag, pol, env
Cont.
Src （v-onc, viral oncogene） probe
RSV-Infected CEF (+ control)
“Normal” chick DNA
癌生物学课程
癌基因与抑癌基因 Oncogene & Tumor Suppressor Gene
提纲
一、癌基因
二、抑癌基因 三、癌基因/抑癌基因与多步骤癌变 四、癌基因/抑癌基因的鉴定
肿瘤发生机理
物理因子 化学因子 生物因子，如病毒
遗传物质改变
正常细胞
癌细胞
➢ a genetic disease characterized by uncontrolled cell growth
Proto-oncogene (c-onc): a normal cellular form of a gene that controls cell
proliferation and can be converted into a cancer-promoting gene by mutation, whose continued activation leads to continued signal transduction, and whose aberrant expression or activity may contribute to tumorigenesis .
D Activated Proto-oncogenes from DNA transfection
Human Bladder Tumor cell line DNA
Isolate high MW DNA
Restriction endonuclease
Isolate DNA fragments
Transfection
B
Transforming (Sarcoma Viruses, RSV)
Peyton Rous, 1911 -cell free lysates could induce sarcomas in other chickens
1，Acute; 2，Transform cultured cells
病毒致癌-- Nobel Prize! 1966
NIH 3T3 fibroblasts
Hale Waihona Puke TransformationIsolate DNA (>99% mouse + 8-10 human genes)
Transformation
Identify Human DNA
Cont.
Use Alu probe Isolate Human DNA
RESULT: A SINGLE human gene is responsible for transforming capability Sequencing
1st human oncogene mutation Ras G12V was identified !
1982, Robert Weinberg, etc three groups
2.癌基因定义 Definition
Viral Oncogene (v-onc): a gene carried by a tumor virus (RNA or DNA), the expression of which is necessary and sufficient to induce transformation in tissue culture cells and tumors in the appropriate animal. V-onc is encoded by cellular sequences that have become inserted into the viral genome.