Mechanisms Involved in Endocytosis of ErbB Proteins
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AbstractCell growth, differentiation and signaling are important processes needed for cell survival. These processes are mediated by binding of growth factors to receptors on the cell membrane. One group of such receptors is the ErbB proteins, which are involved in a wide array of different cellular functions. There are four different ErbB proteins, EGFR, ErbB2, ErbB3 and ErbB4. The ErbB proteins dimerize with and activate each other, leading to activation of intracellular signaling pathways. Downregulation of the receptors from the plasma membrane and a subsequent degradation is an important way to attenuate the signaling. EGFR is known to be downregulated by endocytosis through clathrin-coated pits, and ubiquitination has been suggested to be important for endocytosis. For the other receptors the mechanism of endocytosis is not clear. ErbB proteins have been shown to be related to a range of different cancer types. Dysregulation of ErbB proteins can lead to aberrant signaling and increased cell growth and survival. Knowledge about their regulation is therefore important to be able to target these cancers with specific drugs.
All papers in this work concern endocytosis of the ErbB proteins. ErbB2 is endocytosis resistant, and this ability also affects EGFR in heterodimers with ErbB2. We studied these dimers and found that a reason why ErbB2 inhibits endocytosis of EGFR is that it retains EGFR at the plasma membrane by prohibiting EGF-induced formation of clathrin-coated pits (Paper I). We continued by investigating whether this might also be the case for ErbB3, which is also reported to be endocytosis impaired but we did not find the same correlation. In contrast, we found that endocytosis of ErbB3 was a constitutive process dependent on clathrin (Paper II). We also investigated the endocytosis of ErbB2 itself by inducing its endocytosis with the benzoquinone ansamycin Geldanamycin (GA). Also in this case we found a clear correlation between endocytosis and the nature of the heterodimerization partner. In contrast to cell expressing only ErbB2, cells co-expressing EGFR and/or ErbB3 showed a significant increase in the rate of GA-induced endocytosis of ErbB2 (Paper III). Finally, we investigated the importance of ubiquitin for endocytosis of EGFR and our results support that ubiquitination of EGFR is important for its endocytosis (Paper IV).
Paper I Camilla Haslekås,Kamilla Breen, Ketil W. Pedersen, Lene E. Johannessen, Espen Stang, and Inger Helene Madshus. The inhibitory effect of ErbB2 on epidermal growth factor-induced formation of clathrin-coated pits correlates with retention of epidermal growth factor receptor-ErbB2 oligomeric complexes at the plasma membrane. Molecular Biology of the Cell, Vol. 16, 5832–5842, December 2005. Artikkel
Paper II Kamilla Breen, Sissel B Rønning, Nina Marie Pedersen, Espen Stang and Inger Helene Madshus. ErbB3 is constitutively endocytosed in a clathrin-dependent manner. Manuscript
Paper III Nina Marie Pedersen, Kamilla Breen, Camilla Haslekås, Espen Stang and Inger Helene Madshus. Expression of EGFR or ErbB3 facilitates Geldanamycin-induced downregulation of ErbB2. Submitted
Paper IV Vibeke Bertelsen, Kamilla Breen, Kirsten Sandvig, Espen Stang and Inger Helene Madshus. The Cbl-interacting protein TULA inhibits dynamin-dependent endocytosis. Experimental cell research, Vol.313, 1696-1709, February 2007 Artikkel