PMC:4925210 / 15037-17406 JSONTXT

{"project":"2_test","denotations":[{"id":"27214281-24091015-73835255","span":{"begin":266,"end":267},"obj":"24091015"},{"id":"27214281-24091015-73835255","span":{"begin":266,"end":267},"obj":"24091015"},{"id":"27214281-24091015-73835256","span":{"begin":468,"end":469},"obj":"24091015"},{"id":"27214281-24091015-73835256","span":{"begin":468,"end":469},"obj":"24091015"},{"id":"27214281-19118385-73835257","span":{"begin":1950,"end":1952},"obj":"19118385"},{"id":"27214281-19118385-73835257","span":{"begin":1950,"end":1952},"obj":"19118385"},{"id":"27214281-20107038-73835258","span":{"begin":2001,"end":2003},"obj":"20107038"},{"id":"27214281-20107038-73835258","span":{"begin":2001,"end":2003},"obj":"20107038"}],"text":"WNT9B-induced cell migration is dependent on TRPP2\nWNT5A was shown to promote directional cell migration in a variety of cell types through a process involving both localized Ca2+ release transients originating from the cortical endoplasmic reticulum and Ca2+ influx7. Both types of Ca2+ signaling were required for directional cell migration through the formation of the WNT5A-mediated receptor-actin-myosin polarity structure in the trailing edge of a migrating cell7. Thus, we tested whether WNT9B could induce Ca2+ release. WNT9B did not induce Ca2+ release in wild type or Pkd2-null MEFs, in contrast to ATP which elicited a similar response (Supplementary Fig. 6g,h). Next, we tested whether WNT9B could promote directional cell migration in wild type and Pkd2-null MEFs, through TRPP2 and an increase in intracellular Ca2+ concentration. Wild type or Pkd2-null cells were stimulated with WNT9B for up to 2 h and the number of cells with a typical migratory phenotype (Fig. 7a) was determined in both cultures by double staining for F-actin and Myosin IIB. WNT9B failed to induce this migratory phenotype in Pkd2-null cells, whereas it had a strong effect on wild type cells (Fig. 7a,b). To test whether WNT9B-induced cell polarization was dependent on intracellular Ca2+, Pkd2+/+ MEFs were pretreated with DMSO or 10 μM BAPTA/AM for 45 min before adding WNT9B. BAPTA/AM blocked WNT9B-induced cell polarization (Fig. 7c) to a degree similar to Pkd2-null cells (Fig. 7b). To test whether Pkd2-null MEFs failed to polarize because of TRPP2, we added-back wild type TRPP2 or TRPP2Kv1.3. Wild type, but not TRPP2Kv1.3 rescued cell polarization in Pkd2−/− MEFs (Fig. 7d,e). Consistently, Pkd2-null cells failed to migrate towards a WNT9B gradient using a chemokinetic assay (Fig. 7f,g). In contrast to WNT9B, Pkd2−/− fibroblasts migrated normally in response to PDGF-BB (25 ng/ml) (Fig. 7h), a strong chemokinetic factor acting through Ca2+ flickers41, store-, and/or receptor-activated Ca2+ channels42, indicating that WNT9B-activated PKD1/TRPP2 mediated currents were distinct from the channels activated by PDGF-BB. Overall, these experiments showed that cells lacking TRPP2 exhibited reduced directional migration, suggesting that one of the functions of WNT9B-induced activated TRPP2 in cell culture is to establish polarization during directional cell migration."}