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p120 is a breast tumor suppressor
Our research has shown that dual somatic inactivation of p120 and p53 in the mouse mammary gland leads to high grade and invasive metaplastic carcinoma. While the common denominator of either E-cadherin or p120 inactivation is loss of cell-cell adhesion and metastasis, the phenotypic outcome when ablating p120 is dissimilar. Also, the downstream biochemistry and consequences for the cytoskeleton is different in p120 knockout versus E-cadherin knockout mice. We are trying to understand the biochemical and cellular consequences of p120 loss in breast cancer, which affects 30% of all breast cancer patients. |
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Figure 1: Loss of p120 leads to metaplastic triple negative breast cancer (TNBC) with an inflammatory micro environment.
Shown are p120 positive IDC and p120 negative TNBC. Note the massive influx of CD68 positive macrophages |
p120 controls cytokinesis
We have found that p120 regulates actomyosin contractility at the ingression and cleavage furrow through concomitant binding to RhoA and the centralspindlin component MKLP1, independent of cadherin association.
We have found that p120 regulates actomyosin contractility at the ingression and cleavage furrow through concomitant binding to RhoA and the centralspindlin component MKLP1, independent of cadherin association.
Figure 2:
IF for p120, RhoA and MKLP1 in anaphase and telophase U2OS. Note the co-localization of p120, RhoA and MKLP1 at the equatorial cortex (arrowheads). Right panels show representative magnifications of representative areas denoted by the dotted squares. Bar = 10 μm.
IF for p120, RhoA and MKLP1 in anaphase and telophase U2OS. Note the co-localization of p120, RhoA and MKLP1 at the equatorial cortex (arrowheads). Right panels show representative magnifications of representative areas denoted by the dotted squares. Bar = 10 μm.
Loss of p120 induces multinucleation and chromosomal instability
We observed a high incidence of multinucleated tumour cells and overall severe nuclear atypia in p120-deficient mammary carcinomas from mice and men. To further examine this we made use of breast cancer cell lines observed that loss of p120 leads to overt multi nucleation, demonstrating causality.
We observed a high incidence of multinucleated tumour cells and overall severe nuclear atypia in p120-deficient mammary carcinomas from mice and men. To further examine this we made use of breast cancer cell lines observed that loss of p120 leads to overt multi nucleation, demonstrating causality.
Figure 3:
Control (-dox) and dox-treated (+ dox) mouse and human cancer cell lines transduced with an inducible p120 KD were stained for p120 and Lamin A/ C. Scale bar, 10 mm. Note the overt multi nucleation upon p120 inactivation. dox=doxycyclin
Movie 1:
Shown is a movie of a dividing Trp53∆/∆-p120iKD cell after 3 days on dox. Note the failure to undergo successful completion of cytokinesis after approx. 200 sec. DNA was visualised using H2B-mCherry tagging.
Shown is a movie of a dividing Trp53∆/∆-p120iKD cell after 3 days on dox. Note the failure to undergo successful completion of cytokinesis after approx. 200 sec. DNA was visualised using H2B-mCherry tagging.
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Movie 2:
H2B-mCherry tagged p120iKD U2OS cells undergoing mitosis in the absence (left or top) or presence (right or bottom) of dox. Note the extensive blebbing of the plasma membrane in the p120-deficient cells. |
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Figure 4:
p120 interacts with MKLP1 to regulate focused RhoA activity during cytokinesis IF for p120, RhoA and MKLP1 in dox-treated U2OS p120-iKD cells reconstituted with FL p120-1A (MKLP1 binding) or the p120-1AΔ[1-346] and p120-1AΔ[1-27] truncated (non-MKLP1 binding). Note the broadened RhoA zone in cells reconstituted with p120-1AΔ[1-346] and p120-1AΔ[1-27] (arrowheads). Bar = 10 μm. |
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Translational findings and clinical relevance:
p120 expressed
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p120 lost
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Figure 5:
p120 loss in breast cancer is correlated with decreased disease-specific survival.
Shown are two TMA examples of a tumor expressing p120 (left or top), and a tumor that has lost p120 expression (middle). The findings conform to heterozygous loss of the CTNND1 (p120) allele (not shown). The left (or bottom) graph demonstrates that p120 loss in invasive ductal carcinoma correlates with decreased disease-specific survival (neut and het correspond to the genomic status of CTNND1). Patient data are based on the METABRIC cohort (collaboration Carlos Caldas, Cambridge, UK).
p120 loss in breast cancer is correlated with decreased disease-specific survival.
Shown are two TMA examples of a tumor expressing p120 (left or top), and a tumor that has lost p120 expression (middle). The findings conform to heterozygous loss of the CTNND1 (p120) allele (not shown). The left (or bottom) graph demonstrates that p120 loss in invasive ductal carcinoma correlates with decreased disease-specific survival (neut and het correspond to the genomic status of CTNND1). Patient data are based on the METABRIC cohort (collaboration Carlos Caldas, Cambridge, UK).
Figure 6:
p120, RhoGTPases, Centralspindlin, and cytokinesis regulation.
A During late mitosis, RhoA activity peaks during anaphase and telophase and drives the ingression of the cleavage furrow. Rac1 activity at the polar regions is thought to contribute to cell spreading and adhesion during and/or after cytokinesis. Both positive (Ect2) and negative RhoA regulators (p120 and MP-GAP) localize to the equatorial cortex to allow for rapid GTPase cycling leading to restricted and focused activity of RhoA and subsequent actomyosin contraction. B Recent data confirmed that an inactive RhoA pool (marked by phosphorylation of Ser188; PINK) exists directly adjacent to the active membrane-associated RhoA pool (GREEN) during furrow ingression. The presence of this substantial pool of inactive RhoA will enable rapid GTPase cycling of RhoA to maintain spatially restricted RhoA activity at the equatorial cortex to drive the ingression of the cleavage furrow. The molecular mechanisms underlying phosphorylation of RhoA during cytokineses remain undetermined. Error bar = 2 mm.
C schematic representation of the positioning and function of p120 during anaphase/telophase. MT=microtubule.
p120, RhoGTPases, Centralspindlin, and cytokinesis regulation.
A During late mitosis, RhoA activity peaks during anaphase and telophase and drives the ingression of the cleavage furrow. Rac1 activity at the polar regions is thought to contribute to cell spreading and adhesion during and/or after cytokinesis. Both positive (Ect2) and negative RhoA regulators (p120 and MP-GAP) localize to the equatorial cortex to allow for rapid GTPase cycling leading to restricted and focused activity of RhoA and subsequent actomyosin contraction. B Recent data confirmed that an inactive RhoA pool (marked by phosphorylation of Ser188; PINK) exists directly adjacent to the active membrane-associated RhoA pool (GREEN) during furrow ingression. The presence of this substantial pool of inactive RhoA will enable rapid GTPase cycling of RhoA to maintain spatially restricted RhoA activity at the equatorial cortex to drive the ingression of the cleavage furrow. The molecular mechanisms underlying phosphorylation of RhoA during cytokineses remain undetermined. Error bar = 2 mm.
C schematic representation of the positioning and function of p120 during anaphase/telophase. MT=microtubule.