The AMPK-related kinases NUAK1 and NUAK2 are activated by the tumor suppressor LKB1. We found that NUAK1 interacts with several myosin phosphatases, including the myosin phosphatase targeting-1 (MYPT1)-protein phosphatase-1beta (PP1beta) complex, through conserved Gly-Ile-Leu-Lys motifs that are direct binding sites for PP1beta. Phosphorylation of Ser(445), Ser(472), and Ser(910) of MYPT1 by NUAK1 promoted the interaction of MYPT1 with 14-3-3 adaptor proteins, thereby suppressing phosphatase activity. Cell detachment induced phosphorylation of endogenous MYPT1 by NUAK1, resulting in 14-3-3 binding to MYPT1 and enhanced phosphorylation of myosin light chain-2. Inhibition of the LKB1-NUAK1 pathway impaired cell detachment. Our data indicate that NUAK1 controls cell adhesion and functions as a regulator of myosin phosphatase complexes. Thus, LKB1 can influence the phosphorylation of targets not only through the AMPK family of kinases but also by controlling phosphatase complexes.

EPB41L5 belongs to the band 4.1 superfamily. We investigate here the involvement of EPB41L5 in epithelial–mesenchymal transition (EMT) during mouse gastrulation. EPB41L5 expression is induced during TGFβ-stimulated EMT, whereas silencing of EPB41L5 by siRNA inhibits this transition. In EPB41L5 mutants, cell–cell adhesion is enhanced, and EMT is greatly impaired during gastrulation. Moreover, cell attachment, spreading, and mobility are greatly reduced by EPB41L5 deficiency. Gene transcription regulation during EMT occurs normally at the mRNA level; EPB41L5 siRNA does not affect either the decrease in E-cadherin or the increase in integrin expression. However, at the protein level, the decrease in E-cadherin and increase in integrin are inhibited in both EPB41L5 siRNA-treated NMuMG cells and mutant mesoderm. We find that EPB41L5 binds p120ctn through its N-terminal FERM domain, inhibiting p120ctn–E-cadherin binding. EPB41L5 overexpression causes E-cadherin relocalization into Rab5-positive vesicles in epithelial cells. At the same time, EPB41L5 binds to paxillin through its C terminus, enhancing integrin/paxillin association, thereby stimulating focal adhesion formation.

Embryonic stem cells have the potential to give rise to all cell lineages when introduced into the early embryo. They also give rise to a limited number of different cell types in vitro in specialized culture systems. In this study, we established a culture system in which a structure consisting of lens, neural retina, and pigmented retina was efficiently induced from embryonic stem cells. Refractile cell masses containing lens and neural retina were surrounded by retinal pigment epithelium layers and, thus, designated as eye-like structures. Developmental processes required for eye development appear to proceed in this culture system, because the formation of the eye-like structures depended on the expression of Pax6, a key transcription factor for eye development. The present culture system opens up the possibility of examining early stages of eye development and also of producing cells for use in cellular therapy for various diseases of the eye. Developmental Dynamics 228:664 -671, 2003.

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Here, we report a new serine/threonine protein kinase of the SNF1 subfamily Omphk1. Two Omphk homologues exist in each vertebrate species, and one homologue exists in Drosophila and Caenorhabditis elegans; the kinase domain is highly conserved among these homologues, and several domains are conserved among vertebrate Omphk. Omphk1 expression dynamically changes in the developing central nervous system, is found ubiquitously in epidermis, and is present uniquely in several other tissues. Its expression is also found in each tissue associated with the ventral body wall closure: the primary body wall composed of primitive ectoderm and each component of the secondary body wall. Concomitantly, its null mutant exhibits omphalocele with a failure in closure of the secondary body wall. There are no apparent gross morphological defects in brain, however, despite the unique Omphk1 expression in this tissue.