Hippo pathway and cancer

Question 1

What are the two strands of hippo pathway research?

Answer 1

1. Genes first identified in drosophila screens of tumor repressor genes, when knocked down cause overgrowth (yorki and expanded)
2. Investigation of transcirption factors within muscle promotors (MCATs) with Tead binding to this. YAP then binds to Tead. Hippo binds to YAP linking this together
   These two strands of research were later linked, turning off YAP causes mouse liver overgrowth

Question 2

How do the drosophila and mammalian hippo pathwya differ?

Answer 2

• core kinase casette highly conserved from drosophila
• Yorki in drosophila becomes YAP/TAZ (paralog) in mammals
• Fat receptor in drosophila is GPCR in mammals
• Upstream components vary

Question 3

What is KIBRA?

Answer 3

stands for Kidney and Brain

• Scaffold protein
• No obvious enzyme motifs
• No phosphorylation activity
• Involved with Hippo pathway

Question 4

What is shRNA technology?

Answer 4

• Allows knock down of gene of interest, decraese protein production
• create shRNA which is chopped up by dicer to form siRNAs which is incorporated into the RISC complex and can then cleave complimentary mRNA

Question 5

What is the effect of the knowckdown of KIBRA?

Answer 5

• Decreases phosphorylation of LATS and YAP
• No decrease in MST1/2 or MOB phosphorylation
• Causes change of cells from epithelial to mesenchymal
• Induces expression of mesenchymal markers

Question 6

What is pBabe retroviral expression of genes?

Answer 6

• Plasmid system that produces a virus
• Overexpress gene of interest
• Open reading frame for gene of interest, packaged within a cell line producing virus particles which infect target cells

Question 7

What effect does the overexpression of KIBRA have?

Answer 7

In MCF10A cells already expressing YAP (mesenchymal)

• Increases LATS and YAP phosphorylation
• Antagonizes YAP-induced EMT
• No change in phospho MST (increases with Willin overexpression, must be further up)

Question 8

Does KIBRA have an effect when YAP cannot be phosphorylated?

Answer 8

No

Question 9

Where is it believed that Willin and KIBRA signal?

Answer 9

Willin - phsophorylation of MST
KIBRA - influences phosphorylation of LATS (different to drosophila where KIBRA and willin are believed to be linked, also supported by proteomics)

Question 10

What are the breast cancer subtypes?

Answer 10

• Luminal (ER Positive) treated hormonaly and are the most common
• HER2 are more aggresive, due to genes, treated hormonally and with chemotherapy
• Basal like have no target and are given general chemotherapy and have the worst prognostic category

Question 11

What type of breast cancer does KIBRA play a role in?

Answer 11

• Claudin-low phenotypes of reast cancer have low expression of KIBRA, also shown in patient biopsies
• Shown through hierarchical strucutrin

Question 12

Define an oncogene

Answer 12

Gene that has sustained genetic damage and produces a protein capable of cellular transformation

Question 13

Define a tumor suppressor gene

Answer 13

Class of genes which act in normal cells to inhibit unrestrained cell division and when inactivated place the cell at increased risk of malignant proliferation

Question 14

How do G-protein coupled receptors moduate the Hippo pathway?

Answer 14

• Adding of bovine serum reduces levels of phospho-yap changing where it is located (to nucleus)
• Found that LPA caused decrease of phospho-YAP by itself
• This affects G protein coupled activation, may affect phosphorylation of Lats
• In uveal melanoma mutant Gq/11 promotes tumorgenesis by activating YAP

Question 15

What are the effects of mechanical forces on YAP?

Answer 15

• mechanical stimulus illicit chemical response

- YAP/TAZ is inactivated upon high mechanical force and activated in low mechanical force

Question 16

What is verteporfin?

Answer 16

• Protein which blocks YAP-TEAD interaction
• Used for experiments but possible therapeutic benefit? (approved by FDA)
• blocks pro-YAP oncogenic signals