No abstract

In response to DNA damage, p53 and its homolog p73 have a function antagonistic to NF-B in deciding cell fate. Here, we show for the first time that p73, but not p53, is stabilized by physical interaction with nuclear IB kinase (IKK)-␣ to enhance cisplatin (CDDP)-induced apoptosis. CDDP caused a significant increase in the amounts of nuclear IKK-␣ and p73␣ in human osteosarcoma-derived U2OS cells. Ectopic expression of IKK-␣ prolonged the half-life of p73 by inhibiting its ubiquitination and thereby enhancing its transactivation and pro-apoptotic activities. Consistent with these results, small interfering RNAmediated knockdown of endogenous IKK-␣ inhibited the CDDP-mediated accumulation of p73␣. The kinase-deficient mutant form of IKK-␣ interacted with p73␣, but failed to stabilize it. Furthermore, CDDP-mediated accumulation of endogenous p73␣ was not detected in mouse embryonic fibroblasts (MEFs) prepared from IKK-␣-deficient mice, and CDDP sensitivity was significantly decreased in IKK-␣-deficient MEFs compared with wild-type MEFs. Thus, our results strongly suggest that the nuclear IKK-␣-mediated accumulation of p73␣ is one of the novel molecular mechanisms to induce apoptotic cell death in response to CDDP, which may be particularly important in killing tumor cells with p53 mutation.

The secretory granules of salivary glands often display complex internal substructures, yet little is known of the molecular organization of their contents or the mechanisms involved in packaging of the secretory proteins. We used post-embedding immunogold labeling with antibodies to two secretory proteins, agglutinin and alpha-amylase, to determine their distribution in the Golgi apparatus and secretory granules of the human submandibular gland acinar cells. With monoclonal antibodies specific for carbohydrate epitopes of the agglutinin, reactivity was found in the trans Golgi saccules, trans Golgi network, and immature and mature secretory granules. In the granules, labeling was seen in regions of low and medium electron density, but not in the dense cores. Reactivity seen on the apical and basolateral membranes of acinar and duct cells was attributed to a shared epitope on a membrane glycoprotein. Labeling with a polyclonal antibody to amylase was found in the Golgi saccules, immature and mature secretory granules, but not in the trans Golgi network. In the granules, amylase was present in the dense cores and in areas of medium density, but not in the regions of low density. These results indicate that these two proteins are distributed differently within the secretory granules, and suggest that they follow separate pathways between the Golgi apparatus and forming secretory granules. Small vesicles and tubular structures that labeled only with the antibodies to the agglutinin were observed on both faces of the Golgi apparatus and in the vicinity of the cell membrane. These structures may represent constitutive secretion vesicles involved in transport of the putative membrane glycoprotein to the cell membrane.

No abstract

DNA analyses of 41 individuals with stigmata of Turner syndrome and a 45,X/46,X+mar or 46,X+mar karyotype were carried out. Southern-blot analysis employing 17 Y-specific probes were used to determine whether the marker chromosome was Y-chromosomal in origin. Of the 41 DNA samples from these patients, 23 contained detectable Y-chromosomal DNA. Points of chromosome breakage were distributed over the entire length of the Y long arm. Three individuals, who carry different portions of the Y chromosome, had developed gonadoblastoma. GBY (the gonadoblastoma locus on the Y chromosome) is mapped proximal to DYS132, midway between the 13 Yq loci that we have studied. We also used a polymerase chain reaction technique that could detect 7 loci over the length of the Y chromosome. This technique may be useful for the rapid assessment of marker chromosomes, especially for evaluating the risk of gonadoblastoma.