The perichondrium, a structure made of undifferentiated mesenchymal cells surrounding growth plate cartilage, regulates chondrocyte maturation through poorly understood mechanisms. Analyses of loss- and gain-of-function models show that Twist-1, whose expression in cartilage is restricted to perichondrium, favors chondrocyte maturation in a Runx2-dependent manner. Runx2, in turn, enhances perichondrial expression of Fgf18, a regulator of chondrocyte maturation. Accordingly, compound heterozygous embryos for Runx2 and Fgf18 deletion display the same chondrocyte maturation phenotype as Fgf18-null embryos. This study identifies a transcriptional basis for the inhibition of chondrocyte maturation by perichondrium and reveals that Runx2 fulfills antagonistic functions during chondrogenesis.

Synechococcus elongatus PCC 7942 was the first cyanobacterial strain to be reliably transformed by exogenously added DNA and has become the model organism for cyanobacterial circadian rhythms. With a small genome (2.7 Mb) and well-developed genetic tools, PCC 7942 provides an exceptional opportunity to elucidate the circadian mechanism through genetics. We describe a project to create mutations in every locus of the genome, both to assay each locus for its potential contribution to the circadian clock and to archive data for the cyanobacterial community. Cosmid clones that carry inserts of PCC 7942 DNA are saturated with transposon insertions in vitro to provide sequencing templates and substrates for mutagenesis of the PCC 7942 genome via homologous recombination. We have mutagenized 53% of the chromosome from 50 chromosome-bearing cosmids and identified the positions of insertions in 31 of those cosmids and the 46 kb plasmid, pANL. PCC 7942 mutants defective for 490 different genes have been screened for circadian phenotypes. Mutagenesis of three apparently essential loci, including clpPIIclpX, resulted in circadian phenotypes. We developed an effective antisense suppression method to further the analysis of essential genes. When completed, the set of comprehensive mutations will provide the community with a unique resource whose impact will extend beyond circadian research.

The mammalian constitutive photomorphogenesis 9 (COP9) signalosome (CSN), a protein complex involved in embryonic development, is implicated in cell cycle regulation and the DNA damage response. Its role in tumor development, however, remains unclear. Here, we have shown that the COP9 subunit 6 (CSN6) gene is amplified in human breast cancer specimens, and the CSN6 protein is upregulated in human breast and thyroid tumors. CSN6 expression positively correlated with expression of murine double minute 2 (MDM2), a potent negative regulator of the p53 tumor suppressor. Expression of CSN6 appeared to prevent MDM2 autoubiquitination at lysine 364, resulting in stabilization of MDM2 and degradation of p53. Mice in which Csn6 was deleted died early in embryogenesis (E7.5). Embryos lacking both Csn6 and p53 survived to later in embryonic development (E10.5), which suggests that loss of p53 could partially rescue the effect of loss of Csn6. Mice heterozygous for Csn6 were sensitized to γ-irradiation–induced, p53-dependent apoptosis in both the thymus and the developing CNS. These mice were also less susceptible than wild-type mice to γ-irradiation–induced tumorigenesis. These results suggest that loss of CSN6 enhances p53-mediated tumor suppression in vivo and that CSN6 plays an important role in regulating DNA damage–associated apoptosis and tumorigenesis through control of the MDM2-p53 signaling pathway.