LPA₂ receptor mediates mitogenic signals in human colon cancer cells

Abstract

In the present work, we studied LPA2-mediated signaling using human colon cancer cell lines, which predominantly express LPA2. LPA2 activated Akt and Erk1/2 in response to LPA. LPA mediated Akt activation was inhibited by pertussis toxin (PTX), whereas Erk1/2 activation was completely inhibited by a blocker of phospholipase C␤, U-73122. LPA also induced interleukin-8 (IL-8) synthesis in the colon cancer cells by primarily activating LPA2 receptor. We also found that LPA2 interacts with Na ϩ /H ϩ exchanger regulatory factor 2 (NHERF2). Activation of Akt and Erk1/2 was significantly attenuated by silencing of NHERF2 expression by RNA interference, suggesting a pivotal role of NHERF2 in LPA2-mediated signaling. We found that expression of LPA2 was elevated, whereas expression of LPA1 downregulated in several types of cancers, including ovarian and colon cancer. We conclude that LPA2 is the major LPA receptor in colon cancer cells and cellular signals by LPA2 are largely mediated through its ability to interact with NHERF2.

Lysophosphatidic acid prevents apoptosis of Caco-2 colon cancer cells via activation of mitogen-activated protein kinase and phosphorylation of Bad

“…We have shown previously that Caco-2 cells predominantly express LPA 2 with the expression level of LPA 1 at a significantly lower level and no expression of LPA 3 , making LPA 2 only a significant LPA receptor in these cells [21]. LPA 2 has carboxyl-terminal motifs that allow interaction with a PDZ (PSD-95/DlgA/ZO-1) domain of the Na + /H + exchanger regulatory factor 2 (NHERF2), which functions as a scaffold to link signaling proteins, such as PKA and PKCα, to GPCRs and transporters at the cell surface [21,23,[29][30][31].…”

“…[17][18][19][20][21]. These studies, in conjunction with multiple effects mediated by LPA, suggest that over-expression of LPA 2 may enhance the development of cancer.…”

“…Stable knockdown of NHERF2 in Caco-2 cells using small hairpin RNA (shRNA) targeted against NHERF2 has previously been described [21]. Caco-2 cells with stable knockdown of NHERF2 (Caco-2/CL4) and control transfected cells (Caco-2/ pSiL) were grown in the above media supplemented with 300 unit/ml hygromycin.…”

“…The cell debris was removed by centrifugation at 14,000 g at 4°C for 10 min and protein concentration was determined by the Bicinchoninic Acid assay (Sigma). Equal amounts of lysate were resolved by SDS-PAGE, and Western blot was performed as previously described [21]. Anti-Bad, anti-phospho-Bad, anti-Erk, and anti-phospho-Erk antibodies were purchased from Cell Signaling.…”

“…These scaffold proteins include TRIP6 and at least three PDZ (PSD-95/DlgA/ZO-1) -containing proteins, Na + /H + exchanger regulatory factor 2 (NHERF2), PDZ-RhoGEF, and leukemia-associated RhoGEF [21][22][23][24][25]. We have shown previously that knockdown of NHERF2 in Caco-2 human colon cancer cells significantly attenuated the ability of LPA 2 to induce activation of extracellular signal-regulated kinase-1/2 (Erk), Akt, and interleukin-8 (IL-8) [21]. The consequence of the NHERF2 knockdown was similar to knockdown of LPA 2 as LPA 2 is the major LPA receptor expressed in human colon cancer cells such as Caco-2 cells.…”