Given the widely recognized connection between Golgi and cell migration, it is somehow surprising that studies on Golgi and tumor progression are still very scarce. It is known that most highly invasive tumors have elevated levels of the O-GalNac glycan Tn on the cell surface
83. The presence of glycans at the plasma membrane is known to promote adhesion of cells 84 and could also be involved in the recognition of tumor cells by immune cells 85. Interestingly, tumors with high Tn levels at the plasma membrane show a relocalization of N-acetylgalactosamine-transferases (GalNAc-Ts) from the Golgi to the ER and this causes cells to form more invadopodia and to migrate better 83. Relocalization of several glycosyltransferases was described to happen as a consequence of Golgi fragmentation in prostate cancer 86. Tumors cells often display a fragmented Golgi 87 and we could speculate that Golgi fragmentation is induced to promote cell motility and invasion. However, Golgi fragmentation results in most of the cases in impaired migration of cells in a wound assay, meaning that cells with a fragmented Golgi have a polarity defect. It should be tested whether Golgi fragmentation increases the speed of randomly migrating cells, as it is the case for other conditions where polarity is disrupted. Petrosyan et al. showed that Rab6a associates with
Discussion - 166
NMIIA (non-muscle myosin IIA) and NMIIA prevents Rab6 association with Giantin, necessary to maintain Golgi structure. If NMIIA is depleted, the Golgi becomes compact again, mislocalized glycosyltransferases relocalize to the Golgi and cancer cells become more susceptible to galectin-1 induced apoptosis 86.
Our data confer the Golgi protein GM130 a main role in controlling cell polarity and migration.
We showed that GM130 is progressively lost in colon cancer and often downregulated in breast cancer and thus we speculate an involvement of the Golgi in metastatic spreading.
GM130 depleted cells retain a compact Golgi, but are defective in directional migration.
Despite being not directional, GM130 depleted cells move faster than control cells and become more invasive. GM130 depletion alters two pathways, Cdc42 and Ras. Cdc42 and Ras are both important in cell migration and we could show that inhibition of Ras together with GM130 depletion resulted in a stronger decrease in the ability of the cells to close a wound.
This implies that Cdc42 and Ras affect migration via two different pathways. MDA-MB-231 cells, which have a consitutively active Ras, didn’t migrate faster and didn’t become more invasive after GM130 depletion, whereas cell lines with no mutation in the Ras pathway became faster and more motile upon GM130 depletion. We speculate that a constitutively active Ras might mask the effect of GM130 depletion on cell motility.
How does GM130 control cell velocity? Depletion of the polarity protein Scribble gives phenotypes very similar to the ones observed upon depletion of GM130, thus it is tempting to speculate that GM130 might control Scribble. Scribble connects β-catenin and E-Cadherin and it was shown that the increase in cell velocity caused by loss of Scribble could be dampened by rescuing the interaction between β-catenin and E-Cadherin. GM130 could control Scribble expression levels or alternatively, it could control its localization. It was reported a role for Cdc42 in sorting proteins from the TGN to apical or basolateral membranes
18, therefore we could speculate that by regulating Cdc42 activity at the Golgi, GM130 regulates Scribble positioning. However, it is not known if the same happens when Scribble is mislocalized or if instead it is necessary to lose Scribble to increase cell velocity.
Discussion - 167
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Acknowledgements
The journey that is told in this thesis started 4 years ago and has been one of my most intense periods, with a new research to discover, a new country to explore and new people to know.
My ways crossed with the ways of so many other people that it would take hundreds pages to thank everyone, but everybody I met in this time has somehow contributed to this work.
First of all I want to express my gratitude to my supervisor Dr. Hesso Farhan for of all the energy and time (and money… a lot of money!) that he invested on me. I learnt that the most effective way of teaching is to provide an example and he does exactly this. He showed me how it is to be a scientist but not only this. He contributed to my personal growth by stimulating new interests and by teaching me how to think critically. Thanks to him, after this PhD I have a new vision of science! A work that is also a passion permeating through all the aspects of your life. I feel really lucky because I am one of the few privileged who like their work and this is all merit of Hesso. Together with Hesso, a special thank goes to all the people of the BITg. Veronika, with her infinite patience, Valentina, especially on Thursdays (got it?), Kerstin, Mark, Ricarda, Johanna, Julia and Cristina with whom “I am on the same boat”, literally translated from Italian, an expression with a thousand meanings that can be summarized in: “we walk together, you all know what a PhD is like, we share stressful times as well as parties and this bonds us”. And speaking about Italy, thanks to Nico and Annette who made me feel at home! Without Nico I would probably have died of starvation! Thanks to Dr. Daniel Legler who has been extremely kind with me since when I came for my interview and a special thanks to him because he found the time to be on my thesis committee. Thanks to Josepha, one of the pillars of the BITg, to Ilona, always ready to help, to Edith and Alexandra and to the last arrived Vladimir. Thanks to Conny and Marianne, who make our lab life easier.
Thanks also to Barbara, another positive example during my PhD. Thanks to the Groettrup group at the university, always very available and helpful. An enormous thanks goes also to Dr. Ian Macara, for giving me the opportunity to stay in his lab, where I met new friends like Armelle and Yongliang.
Even if I am work addicted, there are many people living outside the walls of the lab that I have to thank. The Italians Nicola and Nina (now officially Itaian), Paolo, Alessia e Alessio, Giovanni (Losannese!) Ambrogio, Carmelo, Michele, Giorgia (she is probably inside other lab walls) e di nuovo Valentina: è una faticaccia insegnarvi l’italiano, ma d’altronde in qualità di toscano devo diffondere la lingua del sì e le sue parole più importanti come “lapis”,
Acknowledgements - 174
“spengere” e simili! To all the flat mates and friends Carlos, Maria, Kim and Michael and Melanie, with whom I am having a wonderful time at home. When work becomes too much, family and friends are always there to rescue me. Of course, a big thank goes to my parents, who supported me throughout all this time. The “funniest sentence award” goes to my mother who, after a few month I left Italy, discovered the world of Skype and internet and declared: “lo sapevi che col computer si possono fare un sacco di cose?”. And since I wrote in English, she won’t be able to understand! Thanks also to my brothers and the rest of my family, who still don’t know what I do all day, but they think it is cool! Thanks to all my scout friends that share my same pathways even from far away. My old friends Leo, Ale, Andrea, Eli, Bene and the not so old friends Helga and Marco all of them shared several adventures with me during this time. Eli, all the best for the big news of these days! We’ll see each other in Helsinki! Leo, since you are the one who came more often to Konstanz, you win a bonus to come to Paris whenever you like (I like to think that you did not came only for the beer!)! A big thank to the always present (and always late) Filippo, who also embarked himself in the
“spengere” e simili! To all the flat mates and friends Carlos, Maria, Kim and Michael and Melanie, with whom I am having a wonderful time at home. When work becomes too much, family and friends are always there to rescue me. Of course, a big thank goes to my parents, who supported me throughout all this time. The “funniest sentence award” goes to my mother who, after a few month I left Italy, discovered the world of Skype and internet and declared: “lo sapevi che col computer si possono fare un sacco di cose?”. And since I wrote in English, she won’t be able to understand! Thanks also to my brothers and the rest of my family, who still don’t know what I do all day, but they think it is cool! Thanks to all my scout friends that share my same pathways even from far away. My old friends Leo, Ale, Andrea, Eli, Bene and the not so old friends Helga and Marco all of them shared several adventures with me during this time. Eli, all the best for the big news of these days! We’ll see each other in Helsinki! Leo, since you are the one who came more often to Konstanz, you win a bonus to come to Paris whenever you like (I like to think that you did not came only for the beer!)! A big thank to the always present (and always late) Filippo, who also embarked himself in the