Lecture 4: FoxO and Insulin-like receptors Flashcards
(34 cards)
What is the name of the family of transcription factors involved in insulin signalling pathway?
FOXO
Which FOXO family member is expressed at high levels in insulin-responsive tissues?
FOXO1
Describe the structure of FOXO transcription factor DNA binding domain
DNA binding domain consists of three alpha helices, three beta strands, and two wings
- the alpha helices recognise a particular DNA sequence and allow binding of FOXO within the major groove of the DNA strand
- the wings recognise the phosphate backbone
How are FOXO transcription factors regulated?
- phosphorylation by Akt/PKB, CK1 (casein kinase 1) an DYRK1
- acetylation (addition of carboxy group to lysine side chains) by CBP and PCAF
- deacetylation by sirtuins
Describe the main domains of FOXO transcription factors
L1 and L2 = nuclear localisation sequences
E1, E2, and E3 = nuclear export sequences
DNA binding domain
Transactivation domain
How does FOXO phosphorylation result in transcriptional inactivation by insulin signalling?
Insulin signalling activates Akt/PKB, which phosphorylates FOXO
- 14-3-3 dimer recognises and binds to the phosphorylation sites and sterically blocks the nuclear localisation sequences (L1 and L2) and DNA binding domain
- Ran and Crm-1 bind to phosphorylated FoxO resulting in nuclear exclusion
Describe the structure of 14-3-3 proteins
Dimer
each monomer has 9 alpha helices and binds to phosphoserine/phosphotheronine motifs in a sequence specific manner
Describe the movement of FOXO from the cytosol to the nucleus under basal conditions
Importin will bind to NLS along with Ran (GDP form) to promote translocation into the nucleus, DNA binding and gene transcription
Describe the movement of FOXO from the nucleus to the cytosol under basal conditions
Crm1 bind to Ran (GTP form) at nuclear export sequences resulting in FOXO export from the nucleus
Under basal conditions, where is the majority of FOXO located?
Nucleus
Explain how insulin signalling affects nuclear import and export of FOXO
- Akt/PKB phosphorylates FOXO, 14-3-3 binds by recognising phosphoserine/theronine residues, blocks NLS, prevents importin binding and no nuclear import of FOXO
- Akt/PKB phosphorylates FOXO, Ran and Crm1 bind to nuclear export sequences and promotes export to the cytosol
In response to insulin signalling, where is the majority of FOXO located?
cytoplasm
What are some of the roles of FOXO transcription factors?
Inhibit cellular proliferation
promote apoptosis
enhance resistance to oxidative stress
regulate metabolism through their effects in the liver, muscle, adipose tissue and pancreas
increase longevity
(overall action of promoting quiescence, inhibiting growth and cellular proliferation)
How can FOXO gene activation oppose the action of insulin?
in hepatocytes and pancreatic beta cells, FOXO gene activation results in increased blood glucose through decreased insulin secretion
Other than through regulation of FOXO, how else can insulin signalling regulate gene expression?
Activation of the MAPK pathway to stimulate survival, proliferation and differentiation
How does insulin signalling via FOXO and MAPK pathway promote proliferation?
Insulin signalling inhibits FOXO gene activation to prevent apoptosis and promote survival
Insulin signalling via the MAPK pathway to promote expression of genes that improve proliferation and development
Explain how insulin signalling activates the MAPK pathway
- Insulin binds to insulin receptor
- conformational change, transautophosphorylation of the tyrosine kinase domains
- SHC binds to phsophotyrosines on insulin receptor
- SHC phosphorylated by receptor tyrosine kinase
- Grb2 SH2 domains recognise and bind to phosphorylated SHC
- Twin SH3 domains of Grb2 recognise proline rich motifs in SOS protein
- SOS is a guanidine exchange factor for Ras
(small membrane bound GTPase) - SOS promotes exchange of GDP to GTP - activated Ras-GTP activated MAPK phosphorylation pathway (MAPKKK activated by Ras, activates a MAPKK, which then activates the MAPK (E.g. ERK))
- phosphorylated MAPK (E.g. ERK-P) translocates to nucleus and phosphorylates transcription factors that stimulate survival, proliferation and differentiation.
What are similarities and differences between Insulin and Insulin-like growth factors (IGFs)?
unlike insulin, whose C-peptide chain is cleaved form pro-insulin to give the active insulin, the A, B and C peptide chains of IGFs remain attached.
IGFs and insulin are highly homologous (as are their receptors)
How many IGFs are there?
2
(IGF-I and IGF-II)
Where are IGFs mainly produced?
Liver (stimulated by action of growth hormones)
Where do IGFs mainly act?
directly stimulate bone and cartilage growth
True or false: IGFs have a major role in growth regulation prenatally (before birth)
True
What is unique about the expression of IGFs?
There are only expressed on the paternal allele (genetic imprinting - maternal allele silenced - maternally imprinted allele indicates its key role as a regulating factor in foetal growth and development)
What results from forced expression of IGF-I in mouse muscle?
Hypertrophy (size of cells increase rather then number) - much larger mice
