Signal Transduction

Oncogenes

• Activated forms of proto-oncogenes which have a role in proliferation

• Accelerators of cell division

• Over-expressed in cancer

• Gain of function mutations (specific target sites in gene)

Tumour Suppressors

• Negative regulators of proliferation

• Brakes of cell division

• Deleted in cancer

• Loss of function (multiple target sites in the gene)

Signal Transduction

• Important across biology and physiology

• ‘Transfer of extracellular signals to the internal response machinery’

• Signal transduction can tell cells to: divide, stop dividing, apoptose, differentiate, move etc.

• Mitogenic signalling: stimulate cell division

2 examples of oncogenic signalling

1. MAPK - classic ligand driven cancer signalling pathway

2. NF-kB - comparator antiapoptotic pathway

Cell surface receptors growth factor receptors (GFR)

• Epidermal Growth Factor Receptor,

• Platelet Derived GFR,

• Transforming GFR

• Fibroblast GFR

GFR’s or receptor tyrosine kinases (RTK’s)

• Transmembrane proteins with tyrosine kinase activity (>50 different molecules)

• Diverse extracellular domains, conserved intracellular domains (SH2 domains: ‘src homology domains’)

• Internal domain has catalytic properties (phosphorylation and docking sites for protein interaction

The Ras GTPases (G proteins)

• Interact with Receptor Tyrosine Kinases (RTKs) through an intermediate which binds to SH2 domain of RTK

• Binds GTP when active. Needs help to replace GDP for GTP (SOS- son of sevenless)

• When active (GTP-bound) passes on signal to raf kinase

• GTPase activity converts GTP to GDP, inactive

• Ras passes signal (when active) onto Raf kinase to start the phosphorylation cascade

Cytoplasmic signal transduction

• Usually phosphorylation cascade by series of kinases

• Phosphate groups the ‘currency’ used by the cell

• Passed on down the line from kinase to kinase

• Tyrosine kinases, serine/threonine kinases, cAMP kinases

Transcription factors

• MAPK signalling results in activation of ‘ERK’ causing transcription

• These TFs upregulate genes directly, whose proteins subsequently drive cell division (e.g., cell cycle genes)

• E.g., Erk upregulates Cyclin D1 by increasing transcription levels

What are TF’s and their role?

• Ultimate targets of signal transduction

• The ‘internal response machinery’

• The new proteins produced bring about the required change in behaviour

• TF’s are not the only part of the transcription process and require other factors (RNA polymerase etc.)

Control of G-proteins

• GTPase activity of Ras switches off signal

• Therefore one-time signal signal only

• GTPase activity enhanced by GAP proteins (GTPase activating proteins)

• Ras binds GDP more efficiently, needs GEF (SOS) to swap GDP for GTP

• Ras likes to be inactive

Control of signalling

• If kinase activity constant - perpetual mitogenic signalling

• Kinase mediated phosphorylation cascade controlled by corresponding phosphatase enzymes

• TF: Proteolysis prevents accumulation

• TF often dimer, hence co-ordinated by needing 2 subunits

• Other signals can repress transcription. Constant battle between competing signals

Growth Factor Receptors in cancer

• Her-2 (EGFR) often overexpressed in cancers

• Leads to excessive MAPK signalling in those cells

• Results in increased cell proliferation

• Caused by chromosomal amplification of the Her-2 locus, or other means (increased expression)

Herceptin

• First anti-cancer agent produced

• Monoclonal antibody against Her-2

• Blocks excessive signalling in Her-2 positive tumours

• Other EGFR inhibitors now available

• Other Raf and MEK inhibitors available

Ras Oncogene

• Mutated in 50% of colorectal tumours and 70% of pancreatic tumours and others

• Mutations at codons 12, 13 and 61 (GTPase sites) mainly

• Removes the GTPase activity - can’t be switched off

• Results in permanently active MAPK pathway

The NF-kB signalling pathway

• Can be activated in immune cells through ligands . TNF induces the inflammatory response

• Often linked to cancers which start with an inflammatory background

• NF-kB can be activated internally within cells through stress

• In cancer cells, activation can supress apoptosis and encourage cell proliferation and invasion

NF-kB activation

• NFkb a TF, switching on ke cancer related genes

• NFkB is a dimer of 2 out of 5 possible monomers

• Most common dimer (p65 and p50)

• IkB eeps NFkB in the cyctoplasm

• IkB masks the NLS

• IkB phosphorylated by I kappa kinase (IKK) and then degraded

IkB is phosphorylated again, leading to its degradation and activation of NF-kB

Studying signalling errors in cancer

• Transcriptomics (microarray gene expression profiles)

• Proteomics (same as above at protein level)

• Next generation sequencing, find mutations in signalling genes

• More specific approaches pathway by pathway using antibody based approaches (e.g., phospho-antibodies)

• Behaviour based approaches (e.g., invasion/apoptosis)