LKB1 Suppresses p21-activated Kinase-1 (PAK1) by Phosphorylation of Thr109 in the p21-binding Domain

Abstract

The serine/threonine protein kinase LKB1 is a tumor suppressor gene mutated in Peutz-Jeghers syndrome patients. The mutations are found also in several types of sporadic cancer. Although LKB1 is implicated in suppression of cell growth and metastasis, the detailed mechanisms have not yet been elucidated. In this study, we investigated the effect of LKB1 on cell motility, whose acquisition occurs in early metastasis. The knockdown of LKB1 enhanced cell migration and PAK1 activity in human colon cancer HCT116 cells, whereas forced expression of LKB1 in Lkb1-null mouse embryonic fibroblasts suppressed PAK1 activity and PAK1-mediated cell migration simultaneously. Notably, LKB1 directly phosphorylated PAK1 at Thr 109 in the p21-binding domain in vitro. The phosphomimetic T109E mutant showed significantly lower protein kinase activity than wild-type PAK1, suggesting that the phosphorylation at Thr 109 by LKB1 was responsible for suppression of PAK1. Consistently, the nonphosphorylatable T109A mutant was resistant to suppression by LKB1. Furthermore, we found that PAK1 was activated in the hepatocellular carcinomas and the precancerous liver lesions of Lkb1(؉/؊) mice. Taken together, these results suggest that PAK1 is a direct downstream target of LKB1 and plays an essential role in LKB1-induced suppression of cell migration.LKB1 is a serine/threonine kinase whose mutations have been found not only in Peutz-Jeghers syndrome patients (1-3) but also in various types of sporadic cancer (4 -6). These results suggest that LKB1 is a tumor suppressor gene. In addition, we and others (7-9) have previously shown that the heterozygous Lkb1 mutations in mice cause gastrointestinal hamartomas after 20 weeks of age and cause hepatocellular carcinomas (HCCs) 2 after 50 weeks (10). Notably, the Lkb1 gene in all HCC and hepatic precancerous lesions shows loss of heterozygosity (10, 11). These phenotypes in Lkb1(ϩ/Ϫ) mice further indicate that LKB1 is a tumor suppressor gene. In mammalian cells, LKB1 forms a complex with STE20-related adaptor pseudokinase (STRAD) and scaffolding protein MO25, both of which are required for LKB1 enzymatic activity (12, 13). It can phosphorylate and activate at least 14 kinases, including AMP-activated protein kinase (AMPK) and microtubule-associated protein/microtubule affinity-regulating kinases (MARKs) (5). Activation of AMPK by LKB1 leads to inactivation of mammalian target of rapamycin complex 1 via phosphorylation of the tuberous sclerosis complex 1/2, and this pathway has been implicated in tumor suppressor functions of LKB1. In addition to growth control, LKB1 also plays important roles in establishing cell polarity in mammalian cells (14). LKB1 regulates tight junction assembly and cell polarity through AMPK in mammalian cells (15, 16), and we have shown that LKB1 suppresses tubulin polymerization by activating MARK microtubule-associated protein signaling (17). We have also reported that HCCs in Lkb1(ϩ/Ϫ) mice metastasize to the lungs (10). However, the mechanisms by which LKB1 suppresses canc...

Integrin-binding Protein Nischarin Interacts with Tumor Suppressor Liver Kinase B1 (LKB1) to Regulate Cell Migration of Breast Epithelial Cells

“…Thus, these effects might depend on cell type. LKB1 knockdown of human colorectal cells causes an increase in migration and PAK1 activity (23). Loss of epithelial cell polarity has been associated with increased cell invasion (37), and loss of LKB1 has been shown to result in aberrant lung cancer polarity (25), indicating that LKB1 is important in invasion and metastasis through its regulation of cell polarity Our data suggest that suppression of LKB1 and Nischarin enhances cell migration and tumor growth, probably because of defective cell polarity.…”

“…Thus, we used FLAG-tagged deletion constructs of LKB1 ( Cooperative Inhibition of Tumor Cell Migration by Nischarin and LKB1-Our previous work shows that Nischarin inhibits cell migration in several cell types (4,6). Others have shown that LKB1 inhibits migration in esophageal and colorectal cancer cell lines (23,30). Because either LKB1 or Nischarin alone regulates cell migration, we hypothesized that expression of LKB1 and Nischarin together may enhance their ability to inhibit migration.…”

“…LKB1 inhibits PAK1 activation (23). We have shown that Nischarin inhibits PAK1 and LIMK1 activity to inhibit migration and invasion (9, 10).…”

STE20-related Kinase Adaptor Protein α (STRADα) Regulates Cell Polarity and Invasion through PAK1 Signaling in LKB1-null Cells

“…The question of whether an LKB1-independent, STRAD␣-rac1-PAK1 pathway exists when wild-type LKB1 function is intact is difficult to determine since LKB1 itself can alter PAK1 phosphorylation (14,37); thus, discerning the effects of LKB1-STRAD␣-PAK1 from the effects of a separate STRAD␣-PAK1 pathway is difficult with standard approaches. Nevertheless, future studies will attempt to determine if these signaling events unfold in LKB1 wild-type cells and whether LKB1 loss triggers a pro-metastatic STRAD␣ signaling pathway.…”

Effective neurofibromatosis therapeutics blocking the oncogenic kinase PAK1

“…As discussed in detail later, AMPK activates a glucose transport protein called GLUT-4 which is essential for cellular uptake of glucose from blood stream (30). Furthermore, another tumor suppressing kinase called ''LKB1'' activates AMPK, and inactivates PAK1, directly (31). Thus, it is most likely that this tri-terpene blocks PAK1 as well as activates AMPK.…”