Akt Activation Emulates Chk1 Inhibition and Bcl2 Overexpression and Abrogates G2 Cell Cycle Checkpoint by Inhibiting BRCA1 Foci

Akt is perhaps the most frequently activated oncoprotein in human cancers. Overriding cell cycle checkpoint in combination with the inhibition of apoptosis are two principal requirements for predisposition to cancer. Here we show that the activation of Akt is sufficient to promote these two principal processes, by inhibiting Chk1 activation with concomitant inhibition of apoptosis. These activities of Akt cannot be recapitulated by the knockdown of Chk1 alone or by overexpression of Bcl2. Rather the combination of Chk1 knockdown and Bcl2 overexpression is required to recapitulate Akt activities. Akt was shown to directly phosphorylate Chk1. However, we found that Chk1 mutants in the Akt phosphorylation sites behave like wild-type Chk1 in mediating G2 arrest, suggesting that the phosphorylation of Chk1 by Akt is either dispensable for Chk1 activity or insufficient by itself to exert an effect on Chk1 activity. Here we report a new mechanism by which Akt affects G2 cell cycle arrest. We show that Akt inhibits BRCA1 function that induces G2 cell cycle arrest. Akt prevents the translocation of BRCA1 to DNA damage foci and, thereby, inhibiting the activation of Chk1 following DNA damage.     

Influence of genetic architecture on contemporary local evolution in the soapberry bug,Jadera haematoloma: artificial selection on beak length

Little is known about the influence of genetic architecture on local adaptation. We investigated the genetic architecture of the rapid contemporary evolution of mouthparts, the flight polymorphism and life history traits in the soapberry bug Jadera haematoloma (Hemiptera) using laboratory selection. The mouthparts of these seed‐feeding bugs have adapted in 40–50 years by decreasing in length following novel natural selection induced by a host switch to the seeds of an introduced tree with smaller fruits than those of the native host vine. Laboratory selection on beak length in both an ancestral population feeding on the native host and a derived population feeding on the introduced host reveals genetic variance allowing a rapid response (heritabilities of 0.51–0.87) to selection for either longer or shorter beaks. This selection resulted in reverse evolution by restoring long beaks in the derived population and forward evolution by re‐creating short beaks in the ancestral bugs. There were strong genetic correlations (0.68–0.84) in both populations between beak lengths and the frequency of flight morphs, with short beaks associated with short wings. The results reveal a genetically interrelated set of adaptive multivariate traits including both beak length and flight morph. This suite of traits reflects host plant patchiness and seeding phenology. Weaker evidence suggests that egg mass and early egg production may be elements of the same suite. Reversible or forward evolution thus may occur in a broad set of genetically correlated multivariate traits undergoing rapid contemporary adaptation to altered local environments.     

Characterization of a promoter up mutation in the -35 region of the promoter of the primer for ColE1 replication

Summary A point mutation in the -35 region of the promoter of the primer for initiation of DNA replication in the plasmid pMB1 was characterized. This base change causes a promoter up phenotype. The analysis of a second mutant obtained by site-directed mutagenesis allowed the exclusion of a role in the phenotype for the potential intrastrand secondary structure as well as for the methylation state of the DNA in the promoter region. The promoter up phenotype is concluded to be due to a change in the primary structure of the — 35 element with the consequent production of a better cluster of hydrogen bond donors and acceptors for the RNA polymerase.     

Integrated regulation of the photosynthetic gene family from Arabidopsis thaliana in transformed tobacco cells

Summary Expression of the three chlorophyll a/b binding protein (cab) genes of Arabidopsis thaliana was studied in transformed tobacco tissues. For each cab gene, approximately 1000 bp of the promoter region plus a portion of the structural gene was inserted into a promoter-expression vector such that a translational fusion between the cab gene and the promoter-less chloramphenicol acetyltransferase (cat) gene was formed. The constructed molecules were introduced into either cultured tobacco cells or tobacco leaves and the promoter activity was monitored as chloramphenicol acetyltransferase activity. The light-grown tissues exhibited 1.5- to 60-fold greater promoter activity than did dark-grown tissues. Expression of the cab promoters was tissue specific: activities were much stronger in green leaves than other tissues. The cab promoters were almost equally active in transformed calli or shoots derived from leaves. However, in cultured tobacco cells, one promoter was two to three times stronger than the other two. The chimeric gene fusion, cab-cat, segregated in the F1 generation as a dominant Mendelian trait.     

Nucleolus organizer regions and heterochromatin in the zebu (Bos indicus L.)

Summary Ag-NOR staining and a counterstain enhanced fluorescence technique (chromomycin A₃/distamycin A/DAPI-staining = CDD-method) and G-banding, respectively, have been applied to the zebu (Bos indicus L.) chromosomes. The nucleolus organizer regions (NORs) were found in the telomeric regions of chromosomes nos. 2, 3, 4, 11, and 28. CDD staining led to a well-defined R-banding pattern along the chromosome arms and to the visualization of centric heterochromatic bands of variable sizes.     

Genetic variation and distances of five Italian native sheep breeds

Genetic distances among five of the most important Italian native sheep breeds were estimated by using gene frequencies of four blood group and seven blood protein loci and by three different computing methods. The values of the three distance matrices were highly correlated. Genetic distances were found within the range values reported in the literature for local breeds. Shorter distances fitted the historical evidence of migrations.     

Cloning in Bacillus subtilis of temperature-dependent promoter fragments from Bacillus stearothermophilus and Bacillus subtilis

Abstract Using promoter-probe plasmids, more than 200 promoter-containing fragments from Bacillus stearothermophilus and Bacillus subtilis were cloned in B. subtilis . Among these, 15 promoter fragments were highly temperature-dependent in activity compared to the promoter sequence (TTGAAA for the −35 region, TATAAT for the −10 region) of the amylase gene, amyT , from B. stearothermophilus . Some fragments exhibited higher promoter activities at elevated temperature (48°C), others showed higher activities at lower temperature (30°C). Active promoter fragments at higher and lower temperatures were obtained mainly from the thermophile ( B. stearothermophilus ) and the mesophile ( B. subtilis ), respectively. A promoter fragment active at high temperature was sequenced, and the feature of the putative promoter region was discussed.     

Detection of linkage between marker loci and loci affecting quantitative traits in crosses between segregating populations

Summary By making use of pedigree information and information on marker-genotypes of the parent and F-1 individuals crossed to form an F-2 population, it is possible to carry out a linkage analysis between marker loci and loci affecting quantitative traits in a cross between segregating parent populations that are at fixation for alternative alleles at the QTL, but share the same alleles at the marker loci. For two-allele systems, depending on marker allele frequencies in the parent populations, 2–4 times as many F-2 offspring will have to be raised and scored for markers and quantitative traits in order to provide power equivalent to that obtained in a cross between fully inbred lines. Major savings in number of F-2 offspring raised can be achieved by scoring each parent pair for a large number of markers in each chromosomal region and scoring F-1 and F-2 offspring only for those markers for which the parents were homozygous for alternative alleles. For multiple allele systems, particularly when dealing with hypervariable loci, only 10%–20% additional F-2 offspring will have to be raised and scored to provide power equivalent to that obtained in a cross between inbred lines. When a resource population contains novel favorable alleles at quantitative trait loci that are not present (or rare) in a commercial population, analyses of this sort will enable the loci of interest to be identified, mapped and manipulated effectively in breeding programs.Contribution no. 2124-E, 1987 series from The Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel