L3 oncogenes

Question 1

what is the gene aquired by RSV?

Answer 1

Aquires proto-oncogene SRC from host

Question 2

what did collett and erikson do in 1978?

Answer 2

• developed src antiserum (AB to src)
• incubate src immunoprecipitate with radioactive ATP
• lysates from transformed and untransformed cells incubated with specific and non-specific ABS
• found that src is phosphorylating a site on an antibody (band on autoradiogram)
• is src a kinase?

Question 3

what is polyoma largeT?

Answer 3

an oncoprotein - can transform cells

Question 4

what kind of kinase is src?

Answer 4

a tyrosine kinase

Question 5

what are the 3 types of kinases?

Answer 5

serine threonine and tyrosine kinases

Question 6

how was it found that src is a tryosine kinase?

Answer 6

polyoma large t was known to be phosphorylated- aa were separated by chromatography - src lysate added?
- radioactive ATP used - phosphotyrosine dark on gel - auto radiogram
- src must be a tyrosine kinase
(wrongly made buffer which separated phosphorylated tyrosine)

Question 7

what did stan cohen do?

Answer 7

attach EGF to a solid support (in column)

• add lysate from proliferating cells- run through column
• wash and elute
• found a large band that bound to EGF - sequenced - EGF-R
• function map - chemical proteolysis - split into 3 fragments - run on gel with radioactively labelled EGF
• EC fragment bound to EGF

Question 8

what is a kyle and doolittle plot and what does it show about EGF-R?

Answer 8

• it can identify aqueous/lipid loving sequences on protein (dissolve in lipid)
• found the lipid loving sequence on EGF-R was the tmd

Question 9

what was found between src and the protein sequence that bound to EGF?

Answer 9

protein sequence which bound to EGF (EGF-R) was highly similar to src
- src may bind to EGF?

Question 10

how do growth factors induce proliferative effects?

Answer 10

• GF induce cellular effects/proliferation by triggering tyrosine kinase signalling pathways
  (do oncogenes work by triggering signalling in absence of growth factor/EC cues)

Question 11

what did frackleton et al do using EGF?

Answer 11

• incubated cells in a P32 medium
• cell were treated with and without EGF
• digest cells, do chromatography and electrophoresis
• identify phosphoamino-acids
• EGF signalling increased phosphotyrosine
• RTKs are being phosphorylated at PM where/when EGF is added

Question 12

how does the v-ERbB oncogene demonstarte that oncogenes may be working by triggering signalling in the absence of EC cells?

Answer 12

• the v-erbB oncogene is lacking the EC domain

- this means it does not need/respond to EC signals

Question 13

are many growth factor receptors proto-oncogenes?

Answer 13

yes

- are also RTKs

Question 14

give some features of RTK signalling

Answer 14

• when the receptor is off it is in a monomeric state
• when ligand binds oligomerisation/dimerisation occurs
• this results in trans-phosphorylation - dimers phosphorylate one another
• this phosphorylation provides a docking site for downstream signals and the binding of proteins with a RTK domain binding domain.