Meltrin alpha is a member of the metalloprotease-disintegrin (ADAM) family. In this paper we demonstrate that meltrin alpha is involved in the development of white adipose tissue. Compared with wild-type mice, meltrin alpha(-/-) mice displayed moderate resistance to weight gain induced by a high-fat diet, mainly because of an impaired increase in the number of adipocytes. There was no obvious difference in adipocyte size between wild-type and meltrin alpha(-/-) mice, suggesting normal maturation of adipocytes of the latter under a high-fat diet. Embryonic fibroblasts and stromal-vascular cells lacking meltrin alpha exhibited impaired cell proliferation upon adipogenic stimulation, which was accompanied by moderate defects in adipose differentiation. Addition of culture medium conditioned with wild-type cells in an early phase of adipose differentiation did not restore the defects in the meltrin alpha(-/-) cells. These results uncover the involvement of meltrin alpha in the development of obesity and in adipogenic cell proliferation.

Leucine zipper-bearing kinase (LZK) is a novel member of the mixed lineage kinase (MLK) protein family, the cDNA of which was first cloned from a human brain cDNA library [Sakuma, H., Ikeda, A., Oka, S., Kozutsumi, Y., Zanetta, J.-P., and Kawasaki, T. (1997) J. Biol. Chem. 272, 28622-28629]. Several MLK family proteins have been proposed to function as MAP kinase kinase kinases in the c-Jun NH(2) terminal kinase (JNK)/stress-activated protein kinase (SAPK) pathway. In the present study, we demonstrated that, like other MLKs, LZK activated the JNK/SAPK pathway but not the ERK pathway. LZK directly phosphorylated and activated MKK7, one of the two MAPKKs in the JNK/SAPK pathway, to a comparable extent to a constitutive active form of MEKK1 (MEKK1DeltaN), suggesting a biological role of LZK as a MAPKKK in the JNK/SAPK pathway. Recent studies have revealed the essential roles of scaffold proteins in intracellular signaling pathways including MAP kinase pathways. JIP-1, one of the scaffold proteins, has been shown to be associated with MLKs, MKK7, and JNK [Whitmarsh, A.J., Cavanagh, J., Tournier, C., Yasuda, J., and Davis, R.J. (1998) Science 281, 1671-1674], suggesting the presence of a selective signaling pathway including LZK, MKK7, and JNK. Consistent with this hypothesis, we provided evidence that LZK is associated with the C-terminal region of JIP-1 through its kinase catalytic domain. In addition, LZK-induced JNK activation was markedly enhanced when LZK and JNK were co-expressed with JIP-1. These results constituted important clues for understanding the molecular mechanisms regulating the signaling specificities of various JNK activators under different cellular conditions.

Leucine zipper‐bearing kinase (LZK) is a novel member of the mixed lineage kinase (MLK) family [Sakuma, H., Ikeda, A., Oka, S., Kozutsumi, Y., Zanetta, J. P., and Kawasaki, T. (1997) J. Biol. Chem.272, 28622–28629]. We have previously shown that LZK activates the c‐Jun‐NH2 terminal kinase (JNK) pathway, but not the extracellular signal‐related kinase (ERK) pathway, by acting as a mitogen‐activated protein kinase kinase kinase (MAPKKK) [Ikeda, A., Hasegawa, K., Masaki, M., Moriguchi, T., Nishida, E., Kozutsumi, Y., Oka, S., and Kawasaki, T. (2001) J. Biochem.130, 773–781]. However, the mode of activation of LZK remains largely unknown. By means of a yeast two‐hybrid screening system, we have identified a molecule localized to mitochondria, antioxidant protein‐1 (AOP‐1), that binds to LZK and which acts as a modulator of LZK activity. Recently, several MAPKKKs involved in the JNK pathway, such as MEKK1, TAK1 and MLK3, were shown, using over‐expression assay systems, to activate a transcription factor, NF‐κB, through activation of the IKK complex. Using similar assay systems, we demonstrated that LZK activated NF‐κB‐dependent transcription through IKK activation only weakly, but this was reproducible, and that AOP‐1 enhanced the LZK‐induced NF‐κB activation. We also provided evidence that LZK was associated directly with the IKK complex through the kinase domain, and that AOP‐1 was recruited to the IKK complex through the binding to LZK.

The mixed lineage kinase (MLK) family is a recently described protein kinase family. The MLKs contain a kinase domain followed by a dual leucine zipper-like motif. We previously reported the molecular cloning of LZK (leucine zipper-bearing kinase), a novel MLK, and that LZK activated the c-Jun NH 2 terminal kinase (JNK)/stress-activated protein kinase (SAPK) pathway through MKK7 in cells. Here, we reveal that LZK forms dimers/oligomers through its dual leucine zipperlike motif, and that this is necessary for activation of the JNK/ SAPK pathway. We also identify the C-terminal functional region of LZK, which is indispensable for the activation of SEK1, but not that of MKK7. ß

The migration of limb myogenic precursors from limb level somites to their ultimate site of differentiation in the limb is a paradigmatic example of a set of dynamic and orchestrated migratory cell behaviours. The homeobox containing transcription factor ladybird homeobox 1 (Lbx1) is a central regulator of limb myoblast migration, null mutations of Lbx1 result in severe disruptions to limb muscle formation, particularly in the distal region of the limb in mice (Gross et al., 2000). As such Lbx1 has been hypothesized to control lateral migration of myoblasts into the distal limb anlage. It acts as a core regulator of the limb myoblast migration machinery, controlled by Pax3. A secondary role for Lbx1 in the differentiation and commitment of limb musculature has also been proposed (Brohmann et al., 2000; Uchiyama et al., 2000). Here we show that lateral migration, but not differentiation or commitment of limb myoblasts, is controlled by the phosphorylation of three adjacent serine residues of LBX1. Electroporation of limb level somites in the chick embryo with a dephosphomimetic form of Lbx1 results in a specific defect in the lateral migration of limb myoblasts. Although the initial delamination and migration of myoblasts is unaffected, migration into the distal limb bud is severely disrupted. Interestingly, myoblasts undergo normal differentiation independent of their migratory status, suggesting that the differentiation potential of hypaxial muscle is not regulated by the phosphorylation state of LBX1. Furthermore, we show that FGF8 and ERK mediated signal transduction, both critical regulators of the developing limb bud, have the capacity to induce the phosphorylation of LBX1 at these residues. Overall, this suggests a mechanism whereby the phosphorylation of LBX1, potentially through FGF8 and ERK signalling, controls the lateral migration of myoblasts into the distal limb bud.

Leucine zipper-bearing kinase (LZK) is a new member of the mixed lineage protein kinase family. We previously cloned a cDNA encoding LZK from a human cerebellum cDNA library. The following studies indicated that LZK serves as a MAPKKK in the JNK/SAPK pathway in cells, and a scaffold protein, JIP-1, enhances LZK-induced JNK/SAPK pathway activation via physical association. Here we report characterization of the gene structure and fine chromosomal mapping of the human LZK gene. Polymerase chain-reaction (PCR) studies indicated that the human LZK coding sequence is composed of 13 exons, and that all the splice acceptor and donor sequences obey the GT-AG rule. Chromosomal localization studies involving FISH mapping demonstrated that the human LZK gene is located at 3q27.