Abstract
Cdc42-Interacting Protein-4 (CIP4) family adaptors have been implicated in promoting Epidermal Growth Factor Receptor (EGFR) internalization, however, their unique or overlapping functions remain unclear. Here, we show that although CIP4 was not required for early events in clathrin-mediated endocytosis of EGFR, CIP4 localizes to vesicles containing EGFR and Rab5. Furthermore, expression of constitutively active Rab5 led to accumulation of CIP4 and the related adaptor Toca-1 in giant endosomes. Using a mutagenesis approach, we show that localization of CIP4 to endosomes is mediated in part via the curved phosphoinositide-binding face of the CIP4 F-BAR domain. Downregulation of CIP4 in A431 epidermoid carcinoma cells by RNA interference led to elevated EGFR levels, compared to control cells. Although surface expression of EGFR was not affected by CIP4 silencing, EGF-induced transit of EGFR from EEA1-positive endosomes to lysosomes was reduced compared to control cells. This correlated with more robust activation of ERK kinase and entry to S phase in CIP4-depleted A431 cells, compared to control cells. The combined silencing of CIP4 and Toca-1 was more effective in driving cells into S phase, suggesting a partial redundancy in their functions. Overall, our results implicate CIP4 and Toca-1 in regulating late events in EGFR trafficking from endosomes that serves to limit sustained ERK activation within the endosomal compartment.
| Original language | English |
|---|---|
| Pages (from-to) | 1686-1697 |
| Number of pages | 12 |
| Journal | Cellular Signalling |
| Volume | 21 |
| Issue number | 11 |
| DOIs | |
| Publication status | Published - Nov 2009 |
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Keywords
- Adaptor proteins
- EGFR
- F-BAR domain
- Rab5
- Vesicle trafficking
ASJC Scopus subject areas
- Cell Biology
Cite this
F-BAR-containing adaptor CIP4 localizes to early endosomes and regulates Epidermal Growth Factor Receptor trafficking and downregulation. / Hu, Jinghui; Troglio, Flavia; Mukhopadhyay, Alka; Everingham, Stephanie; Kwok, Ester; Scita, Giorgio; Craig, Andrew W B.
In: Cellular Signalling, Vol. 21, No. 11, 11.2009, p. 1686-1697.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - F-BAR-containing adaptor CIP4 localizes to early endosomes and regulates Epidermal Growth Factor Receptor trafficking and downregulation
AU - Hu, Jinghui
AU - Troglio, Flavia
AU - Mukhopadhyay, Alka
AU - Everingham, Stephanie
AU - Kwok, Ester
AU - Scita, Giorgio
AU - Craig, Andrew W B
PY - 2009/11
Y1 - 2009/11
N2 - Cdc42-Interacting Protein-4 (CIP4) family adaptors have been implicated in promoting Epidermal Growth Factor Receptor (EGFR) internalization, however, their unique or overlapping functions remain unclear. Here, we show that although CIP4 was not required for early events in clathrin-mediated endocytosis of EGFR, CIP4 localizes to vesicles containing EGFR and Rab5. Furthermore, expression of constitutively active Rab5 led to accumulation of CIP4 and the related adaptor Toca-1 in giant endosomes. Using a mutagenesis approach, we show that localization of CIP4 to endosomes is mediated in part via the curved phosphoinositide-binding face of the CIP4 F-BAR domain. Downregulation of CIP4 in A431 epidermoid carcinoma cells by RNA interference led to elevated EGFR levels, compared to control cells. Although surface expression of EGFR was not affected by CIP4 silencing, EGF-induced transit of EGFR from EEA1-positive endosomes to lysosomes was reduced compared to control cells. This correlated with more robust activation of ERK kinase and entry to S phase in CIP4-depleted A431 cells, compared to control cells. The combined silencing of CIP4 and Toca-1 was more effective in driving cells into S phase, suggesting a partial redundancy in their functions. Overall, our results implicate CIP4 and Toca-1 in regulating late events in EGFR trafficking from endosomes that serves to limit sustained ERK activation within the endosomal compartment.
AB - Cdc42-Interacting Protein-4 (CIP4) family adaptors have been implicated in promoting Epidermal Growth Factor Receptor (EGFR) internalization, however, their unique or overlapping functions remain unclear. Here, we show that although CIP4 was not required for early events in clathrin-mediated endocytosis of EGFR, CIP4 localizes to vesicles containing EGFR and Rab5. Furthermore, expression of constitutively active Rab5 led to accumulation of CIP4 and the related adaptor Toca-1 in giant endosomes. Using a mutagenesis approach, we show that localization of CIP4 to endosomes is mediated in part via the curved phosphoinositide-binding face of the CIP4 F-BAR domain. Downregulation of CIP4 in A431 epidermoid carcinoma cells by RNA interference led to elevated EGFR levels, compared to control cells. Although surface expression of EGFR was not affected by CIP4 silencing, EGF-induced transit of EGFR from EEA1-positive endosomes to lysosomes was reduced compared to control cells. This correlated with more robust activation of ERK kinase and entry to S phase in CIP4-depleted A431 cells, compared to control cells. The combined silencing of CIP4 and Toca-1 was more effective in driving cells into S phase, suggesting a partial redundancy in their functions. Overall, our results implicate CIP4 and Toca-1 in regulating late events in EGFR trafficking from endosomes that serves to limit sustained ERK activation within the endosomal compartment.
KW - Adaptor proteins
KW - EGFR
KW - F-BAR domain
KW - Rab5
KW - Vesicle trafficking
UR - http://www.scopus.com/inward/record.url?scp=69249244189&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=69249244189&partnerID=8YFLogxK
U2 - 10.1016/j.cellsig.2009.07.007
DO - 10.1016/j.cellsig.2009.07.007
M3 - Article
VL - 21
SP - 1686
EP - 1697
JO - Cellular Signalling
JF - Cellular Signalling
SN - 0898-6568
IS - 11
ER -