Functional cooperation among Ras, STAT5, and phosphatidylinositol 3-kinase is required for full oncogenic activities of BCR/ABL in K562 cells.

BCR/ABL tyrosine kinase generated from the chromosomal translocation t(9;22) causes chronic myelogenous leukemia and acute lymphoblastic leukemia. To examine the roles of BCR/ABL-activated individual signaling molecules and their cooperation in leukemogenesis, we inducibly expressed a dominant negative (DN) form of Ras, phosphatidylinositol 3-kinase, and STAT5 alone or in combination in p210 BCR/ABL-positive K562 cells. The inducibly expressed DN Ras (N17), STAT5 (694F), and DN phosphatidylinositol 3-kinase (Delta p85) inhibited the growth by 90, 55, and 40%, respectively. During the growth inhibition, the expression of cyclin D2 and cyclin D3 was suppressed by N17, 694F, or Delta p85; that of cyclin E by N17; and that of cyclin A by Delta p85. In addition, N17 induced apoptosis in a small proportion of K562, whereas 694F and Delta p85 were hardly effective. In contrast, coexpression of two DN mutants in any combinations induced severe apoptosis. During these cultures, the expression of Bcl-2 was suppressed by N17, 694F, or Delta p85, and that of Bcl-XL by N17. Furthermore, although K562 was resistant to interferon-alpha- and dexamethasone-induced apoptosis, disruption of one pathway by N17, 694F, or Delta p85 sensitized K562 to these reagents. These results suggested that cooperation among these molecules is required for full leukemogenic activities of BCR/ABL.     

Comparative potencies of botulinum toxin for infant and adult mice

Culture fluids of 4 type A, 6 type B, and 1 type EClostridium botulinum strains were compared for their toxicities in 9–11-day-old (infant) and adult mice. In intraperitoneal challenges, most toxins were about equally potent for infants and adults; the exceptions were 4 type B preparations, which were 500 to 5,000 times more lethal for infants. In orogastric challenges, samples were 10 to >15,000 times more toxic for infants, and the previously mentioned type B toxins constituted a group having the highest relative toxicity for infants.     

A RelA-SpoT homolog (Cr-RSH) identified in Chlamydomonas reinhardtii generates stringent factor in vivo and localizes to chloroplasts in vitro

A gene encoding a putative guanosine 3′,5′-bispyrophosphate (ppGpp) synthase–degradase, designated Cr-RSH, was identified in the unicellular photosynthetic eukaryote Chlamydomonas reinhardtii. The encoded Cr-RSH protein possesses a putative chloroplast-targeting signal at its NH₂-terminus, and translocation of Cr-RSH into chloroplasts isolated from C.reinhardtii was demonstrated in vitro. The predicted mature region of Cr-RSH exhibits marked similarity to eubacterial members of the RelA–SpoT family of proteins. Expression of an NH₂-terminal portion of Cr-RSH containing the putative ppGpp synthase domain in a relA, spoT double mutant of Escherichia coli complemented the growth deficits of the mutant cells. Chromatographic analysis of ³²P-labeled cellular mononucleotides also revealed that expression of Cr-RSH in the mutant bacterial cells resulted in the synthesis of ppGpp. SpoT, which catalyzes (p)ppGpp degradation, is dispensable in E.coli only if cells also lack RelA, which possesses (p)ppGpp synthase activity. The complementation analysis thus indicated that Cr-RSH possesses both ppGpp synthase and degradase activities. These results represent the first demonstration of ppGpp synthase–degradase activities in a eukaryotic organism, and they suggest that eubacterial stringent control mediated by ppGpp has been conserved during evolution of the chloroplast from a photosynthetic bacterial symbiont.     

Bcl-2 inhibits apoptosis of spermatogonia and growth of spermatogonial stem cells in a cell-intrinsic manner

The growth, differentiation, and death/survival of spermatogonia are precisely regulated for the proper production of spermatozoa. We have previously shown that Bcl-2 ectopically expressed in spermatogonia caused the inhibition of normal spermatogonial apoptosis and the subsequent failure of differentiation in transgenic mice. In addition, the growth of spermatogonial stem cells seemed to be temporally arrested in the transgenic mice. In the present study, we attempted to examine whether the abnormality of spermatogonia described above was caused by Bcl-2 misexpression in the spermatogonia or by an abnormal spermatogenic environment of the transgenic mice. We transplanted testicular cells of transgenic mice to seminiferous tubules of W/Wv mice in which transplanted normal testicular cells can undergo spermatogenesis. We found that the transplanted spermatogonia of the transgenic mice reproduced a series of abnormal changes including temporal growth arrest of spermatogonial stem cells and abnormal accumulation of spermatogonia in tubules, which were also observed in the testes of the transgenic mice. The results indicated that Bcl-2 inhibited apoptosis of spermatogonia and growth of spermatogonial stem cells in a cell-intrinsic manner. We also cultured testicular cells of transgenic mice and found that the spermatogonia of the transgenic mice were better able to survive than were those of wild-type mice but that their differentiation was not affected. The result suggested that failure of differentiation of the accumulated spermatogonia in the transgenic testes is not due to the abnormality of the bcl-2 misexpressing spermatogonia, but may be caused by extrinsic problems including improper interaction of spermatogonia with supporting cells.     

Late Paleozoic faunal, climatic, and geographic changes in the Baoshan block as a Gondwana-derived continental fragment in southwest China

The Carboniferous and Permian of the Baoshan block consist of three major depositional sequences: a Lower Carboniferous carbonate sequence, a Lower Permian siliciclastic sequence, and a Middle Permian carbonate sequence. These three sequences were interrupted by two major regressive events: first, the Namurian Uplift ranging in age from Serpukovian to Gzhelian, and second, the Post-Sakmarian Regression occurring probably at Artinskian time in the Baoshan block, although the precise time interval of the latter event is still unclear. The Baoshan block is characterized by warm-water, highly diverse and abundant faunas during the Early Carboniferous, by cold-water and low diversity faunas during the Early Permian, and by possibly warm-water but low diversity faunas during the Middle Permian. The Sweetognathus bucaramangus conodont fauna constrains the upper boundary of the diamictite-bearing siliciclastic deposits (Dingjiazhai Formation) to the Sakmarian to early Artinskian, as well as the eruption of the rifting basalts (Woniusi Formation) to, at least, the post-early Artinskian. Paleozoogeographically, affiliation of the faunas in the Baoshan block changed from Eurasian in the Early Carboniferous, to Peri-Gondwanan in the Early Permian, and to Marginal Cathaysian/Cimmerian in the Middle Permian. Cimmerian blocks have more or less comparable geohistory to one another in the Carboniferous and Permian. During the Middle Permian, the eastern Cimmerian blocks such as Sibumasu (s.s), Baoshan, and Tengchong are not far from the palaeoequator, but apparently more distant than the western Cimmerian blocks based on the presence or absence of some index taxa such as the fusulinaceans Eopolydiexodina and Neoschwagerina, and the corals Thomasiphyllum and Wentzellophyllum persicum.     

Behavior of Psychrotrophic Lactic Acid Bacteria Isolated from Spoiling Cooked Meat Products

Three kinds of lactic acid bacteria were isolated from spoiling cooked meat products stored below 10°C. They were identified as Leuconostoc mesenteroides subsp. mesenteroides, Lactococcus lactis subsp. lactis, and Leuconostoc citreum. All three strains grew well in MRS broth at 10°C. In particular, L. mesenteroides subsp. mesenteroides and L. citreum grew even at 4°C, and their doubling times were 23.6 and 51.5 h, respectively. On the other hand, although the bacteria were initially below the detection limit (<10 CFU/g) in model cooked meat products, the bacterial counts increased to 10⁸ CFU/g at 10°C after 7 to 12 days.     

Role of the loop structure of the catalytic domain in rice class I chitinase

In the three-dimensional structure of a rice class I chitinase (OsChia1b) determined recently, a loop structure (loop II) is located at the end of the substrate-binding cleft, and is thus suggested to be involved in substrate binding. In order to test this assumption, deletion of the loop II region from the catalytic domain of OsChia1b and replacement of Trp159 in loop II with Ala were carried out. The loop II deletion and the W159A mutation increased hydrolytic activity not only towards (GlcNAc)6 but also towards polysaccharide substrates. Similar results were obtained for kcat/Km values determined for substrate reduced-(GlcNAc)5. The two mutations shifted the splitting positions in (GlcNAc)6 to the reducing end side, but the shift was less intensive in the Trp mutant. Theoretical analysis of the reaction time course indicated that sugar residue affinity at the +3 subsite was reduced from -2 kcal/mol to +0.5 kcal/mol by loop II deletion. Reduced affinity at the +3 subsite might enhance the release of product fragments, resulting in higher turnover and higher enzymatic activities. Thus, we concluded that loop II is involved in sugar residue binding at the +3 subsite, but that Trp159 itself appears to contribute only partly to sugar residue interaction at the subsite.