Mitogen-activated protein kinase (MAPK) is a conserved eukaryotic signaling factor that mediates various signals, cumulating in the activation of transcription factors. Extracellular signal-regulated kinase (ERK), a M...
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Mitogen-activated protein kinase (MAPK) is a conserved eukaryotic signaling factor that mediates various signals, cumulating in the activation of transcription factors. Extracellular signal-regulated kinase (ERK), a MAPK? is activated through phosphorylation by the kinase MAP/ERK kinase (MEK). To elucidate the extent of the involvement of ERR in various aspects of animal development, we searched for a Drosophila mutant which responds to elevated MEK activity and herein identified a lace mutant. Mutants with mild lace alleles grow to become adults with multiple aberrant morphologies in the appendages, compound eye, and bristles. These aberrations were suppressed by elevated MEK activity. Structural and transgenic analyses of the lace cDNA have revealed that the lace gene product is a membrane protein similar to the yeast protein LcB2, a subunit of serine palmitoyltransferase (SPT), which catalyzes the first step of sphingolipid biosynthesis. In fact, SPT activity in the fly expressing epitope-tagged Lace was absorbed by epitope-specific antibody. The number of dead cells in various imaginal discs of a lace hypomorph was considerably increased, thereby ectopically activating c-Jun N-terminal kinase (JNK), another MAPK These results account for the adult phenotypes of the lace mutant and suppression of the phenotypes by elevated MEK activity we hypothesize that mutation of lace causes decreased de novo synthesis of sphingolipid metabolites, some of which are signaling molecules, and one or more of these changes activates JNK to elicit apoptosis, The ERK pathway may be antagonistic to the JNK pathway in the control of cell survival.
BAcKGROUND: Myriocin is a natural product that potently induces apoptosis of a murine cytotoxic T lymphocyte cell line (cTLL-2) and inhibits a serine palmitoyltransferase (SPT) activity that has been detected in cell ...
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BAcKGROUND: Myriocin is a natural product that potently induces apoptosis of a murine cytotoxic T lymphocyte cell line (cTLL-2) and inhibits a serine palmitoyltransferase (SPT) activity that has been detected in cell extracts and is thought to initiate sphingolipid biosynthesis. Because SPT has never been biochemically purified and a comprehensive appraisal of myriocin-binding proteins has not been conducted, we isolated specific targets using myriocin-based affinity chromatography. RESULTS: Myriocin derivatives were synthesized and evaluated using cTLL-2 proliferation and SPT activity assays. Guided by these results, affinity chromatography matrices were prepared and two specific myriocin-binding proteins were isolated from cTLL-2 lysates. Analyses of these polypeptides establish conclusively that they are murine LcB1 and LcB2, mammalian homologs of two yeast proteins that have been genetically linked to sphingolipid biosynthesis. cONcLUSION: The ability of myriocin-containing matrices to bind factors that have SPT activity and the exclusive isolation of LcB1 and LcB2 as myriocin-binding proteins demonstrates that the two proteins are directly responsible for SPT activity and that myriocin acts directly upon these polypeptides.
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