Functional interaction between mouse erbB3 and wild-type rat c-neu in transgenic mouse mammary tumor cells.
BACKGROUND
Co-expression of several receptor tyrosine kinases (RTKs), including erbB2 and erbB3, is frequently identified in breast cancers. A member of the RTK family, the kinase-deficient erbB3 can activate downstream signaling via heterodimer formation with erbB2. We studied the expression of RTK receptors in mammary tumors from the wild-type (wt) rat c-neu transgenic model. We hypothesized that physical and functional interactions between the wt rat neu/ErbB2 transgene and mouse ErbB3-encoded proteins could occur, activating downstream signaling and promoting mammary oncogenesis.


METHODS
Immunohistochemical and Western blot analyses were performed to study the expression of rat c-neu/ErbB2 and mouse erbB3 in mammary tumors and tumor-derived cell lines from the wt rat c-neu transgenic mice. Co-immunoprecipitation methods were employed to quantitate heterodimerization between the transgene-encoded protein erbB2 and the endogenous mouse erbB3. Tumor cell growth in response to growth factors, such as Heregulin (HRG), epidermal growth factor (EGF), or insulin-like growth factor-1 (IGF-1), was also studied. Post-HRG stimulation, activation of the RTK downstream signaling was determined by Western blot analyses using antibodies against phosphorylated Akt and mitogen-activated protein kinase (MAPK), respectively. Specific inhibitors were then used with cell proliferation assays to study the phosphoinositide-3 kinase (PI-3K)/Akt and MAPK kinase (MEK)/MAPK pathways as possible mechanisms of HRG-induced tumor cell proliferation.


RESULTS
Mammary tumors and tumor-derived cell lines frequently exhibited elevated co-expression of erbB2 and erbB3. The transgene-encoded protein erbB2 formed a stable heterodimer complex with endogenous mouse erbB3. HRG stimulation promoted physical and functional erbB2/erbB3 interactions and tumor cell growth, whereas no response to EGF or IGF-1 was observed. HRG treatment activated both the Akt and MAPK pathways in a dose- and time-dependent manner. Both the PI-3K inhibitor LY 294002 and MEK inhibitor PD 98059 significantly decreased the stimulatory effect of HRG on tumor cell proliferation.


CONCLUSIONS
The co-expression of wt rat neu/ErbB2 transgene and mouse ErbB3, with physical and functional interactions between these two species of RTK receptors, was demonstrated. These data strongly suggest a role for erbB3 in c-neu (ErbB2)-associated mammary tumorigenesis, as has been reported in human breast cancers.
Kim
Aeree
A
Department of Pathology and College of Medicine, Oklahoma University Health Sciences Center (OUHSC), Oklahoma City, OK, USA. ark@korea.ac.kr
Liu
Bolin
B
Ordonez-Ercan
Dalia
D
Alvarez
Kathy M
KM
Jones
Lynn D
LD
McKimmey
Christine
C
Edgerton
Susan M
SM
Yang
Xiaohe
X
Thor
Ann D
AD
D000818:Animals; D001943:Breast Neoplasms; D002455:Cell Division; D045744:Cell Line, Tumor; D004273:DNA, Neoplasm / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D006023:Glycoproteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D015674:Mammary Neoplasms, Animal / Q000235:genetics* / Q000473:pathology; D051379:Mice; D008822:Mice, Transgenic; D051381:Rats; D018719:Erb-b2 Receptor Tyrosine Kinases; D020893:Receptor, ErbB-3 / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D004273:DNA, Neoplasm; D006023:Glycoproteins; D018719:Erb-b2 Receptor Tyrosine Kinases; D020893:Receptor, ErbB-3; C483804:Erbb2 protein, rat