Maternal Wnt/STOP signaling promotes cell division during early Xenopus embryogenesis.

During Xenopus development, Wnt signaling is thought to function first after midblastula transition to regulate axial patterning via ß-catenin-mediated transcription. Here, we report that Wnt/glycogen synthase kinase 3 (GSK3) signaling functions posttranscriptionally already in mature oocytes via Wnt/stabilization of proteins (STOP) signaling. Wnt signaling is induced in oocytes after their entry into meiotic metaphase II and declines again upon exit into interphase. Wnt signaling inhibits Gsk3 and thereby protects proteins from polyubiquitination and degradation in mature oocytes. In a protein array screen, we identify a cluster of mitotic effector proteins that are polyubiquitinated in a Gsk3-dependent manner in Xenopus. Consequently inhibition of maternal Wnt/STOP signaling, but not ß-catenin signaling, leads to early cleavage arrest after fertilization. The results support a novel role for Wnt signaling in cell cycle progression independent of ß-catenin.

One step cloning of defined DNA fragments from large genomic clones.

Recently, the nucleotide sequences of entire genomes became available. This information combined with older sequencing data discloses the exact chromosomal location of millions of nucleotide markers stored in the databases at NCBI, EMBO or DDBJ. Despite having resolved the intron/exon structures of all described genes within these genomes with a stroke of a pen, the sequencing data opens up other interesting possibilities. For example, the genomic mapping of the end sequences of the human, murine and rat BAC libraries generated at The Institute for Genomic Research (TIGR), reveals now the entire encompassed sequence of the inserts for more than a million of these clones. Since these clones are individually stored, they are now an invaluable source for experiments which depend on genomic DNA. Isolation of smaller fragments from such clones with standard methods is a time consuming process. We describe here a reliable one-step cloning technique to obtain a DNA fragment with a defined size and sequence from larger genomic clones in less than 48 hours using a standard vector with a multiple cloning site, and common restriction enzymes and equipment. The only prerequisites are the sequences of ends of the insert and of the underlying genome.