It is well known that extracellular pH (pH
e) tumor tissue is acidic, and sometimes induces matrix metalloproteinase-9 (MMP-9) expression, invasion and metastasis. The small G protein RhoA plays a role in the intracellular signaling pathway for acidic pH
e. In this study, PAK6 and PAK7, two of the p21 activated kinases (PAKs) that are known to act downstream of small G proteins, were investigated and their role in acidic pH
e signaling inducing MMP-9 expression was examined. PAK6 and PAK7 double knockout cell variants (PAK6/7 KO cells) of mouse B16-BL6 melanoma were prepared via the CRISPR / Cas9 method. MMP-9 activity and mRNA levels were analyzed by gelatin zymography and quantitative PCR, respectively. Enforced expression of PAK6/7 decreased the induction of MMP-9 in parental B16-BL6 cells. On the contrary, the acidic pH
e-induced MMP-9 expression level in PAK6/7 KO cells was higher than in parental cells, while MMP-9 induction was absent in PAK6/7 KO cells at neutral pH
e. When PAK6/7 KO cells were rescued by introduction of vectors expressing wild type PAK6/7 and its mutants including constitutively active and kinasedead forms, acidic pH
e-induced MMP-9 expression was suppressed by the introduction of wild-type and constitutively active PAK6/7, but not by the kinase-dead forms. These results suggest that PAK6/7 play a role of negative feedback in acidic pH
e signaling to induce MMP-9 expression.
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