Direct Evidence for Changes in the Resistance of Legume Root Nodules to O2 Diffusion

Indirect evidence suggests that legumes can adjust rapidly theresistance of their root nodules to O₂ diffusion. Here we describeexperiments using O₂ specific micro-electrodes and dark fieldmicroscopy to study directly the operation of this diffusionbarrier. The O₂ concentration sensed by the electrode decreasedsharply in the region of the inner cortex and was less than1.0 mmol m^–3 throughout the infected tissue in nodulesof both pea (Pisum sativum) and french bean (Phaseolus vulgaris).In a number of experiments the ambient O₂ concentration wasincreased to 40% while the electrode tip was just inside theinner cortex. In 13 out of 21 cases the O₂ concentration atthis position either remained low and unchanged or increasedirreversibly to near ambient values. In the remaining casesthe O₂ concentration increased after 1 to 2.5 min and then decreasedto its former value. These results are ascribed to an increasein resistance of the barrier in response to increased O₂ fluxinto the nodule. It was shown microscopically that air spacesboth at the boundary between the infected zone and the innercortex, and within the infected zone started to disappear 3min after nodules were exposed to high ambient O₂ concentrationsand had disappeared completely after 8 min. These spaces werenot changed by exposure of the nodule for 10 min to either N₂or air. Key words: Oxygen, root nodules, air spaces     

Evolution of the 28S ribosomal RNA gene in anurans: regions of variability and their phylogenetic implications

Fifteen restriction sites were mapped to the 28S ribosomal RNA gene of individuals representing 54 species of frogs, two species of salamanders, a caecilian, and a lungfish. Eight of these sites were present in all species examined, and two were found in all but one species. Alignment of these conserved restriction sites revealed, among anuran 28S rRNA genes, five regions of major length variation that correspond to four of 12 previously identified divergent domains of this gene. One of the divergent domains (DD8) consists of two regions of length variation separated by a short segment that is conserved at least throughout tetrapods. Most of the insertions, deletions, and restriction-site variations identified in the 28S gene will require sequence-level analysis for a detailed reconstruction of their history. However, an insertion in DD9 that is coextensive with frogs in the suborder Neobatrachia, a BstEII site that is limited to representatives of two leptodactylid subfamilies, and a deletion in DD10 that is found only in three ranoid genera are probably synapomorphies.      

Dissecting the activating mutations in v-erbB of avian erythroblastosis virus strain R.

The v-erbB oncogene isolated from the R (or ES4) strain of avian erythroblastosis virus is capable of inducing erythroleukemia and fibrosarcomas. This oncogene differs from the proto-oncogene c-erbB, the avian homolog of the epidermal growth factor receptor, by its lack of an intact ligand-binding domain as well as additional alterations in its cytoplasmic coding sequences. By contrast, the insertionally activated c-erbB, a variant oncogene, which encodes a product that also lacks the ligand-binding domain but is otherwise unaltered in its cytoplasmic coding sequences, is capable of inducing leukemia but cannot induce sarcomas. In this report, we show that the critical changes for activating the sarcomagenic potential displayed by v-erbB R are two point mutations within the tyrosine kinase domain and an internal deletion of 21 amino acids in the carboxyl-terminal regulatory domain. The removal of the carboxyl-terminal autophosphorylation sites is not obligatory. These activating mutations (Arg-263 to His, Ile-384 to Ser, and the deletion of residues 494 to 514), when introduced singly into the insertionally activated c-erbB, all dramatically increase fibroblast-transforming potential. Arg-263 resides near the highly conserved HRD motif of the kinase domain, and its mutation to His increases the autophosphorylation activity. The other two mutations do not alter the intrinsic kinase activity and presumably affect other aspects of the receptor involved in growth signaling. Therefore, the high transforming potential of v-erbB R is a consequence of synergism among multiple activating mutations.     

Differential behaviour of 25-hydroxyvitamin D3 24-hydroxylase and 1-hydroxylase in response to protein synthesis inhibitors and cytochrome P-450 inhibitors

To characterize 25-hydroxyvitamin D3 24-hydroxylase and 25-hydroxyvitamin D3 1-hydroxylase, the activities of the two enzymes were measured in the presence of two types of inhibitors. The effect of protein synthesis inhibitors on 25-hydroxyvitamin D3-stimulated 24-hydroxylase activity in 1-hydroxylating rat kidneys perfused in vitro was tested. Actinomycin D (4 microM) and cycloheximide (10 microM) each abolished 25-hydroxyvitamin D3 24-hydroxylase synthesis when added at the start of perfusion but not when added 4 h later; they did not affect 25-hydroxyvitamin D3 1-hydroxylase activity. The effects of cytochrome P-450 inhibitors on the two enzyme activities were then studied in vivo. Metyrapone and SKF-525A (50 mg/kg body weight) each inhibited 25-hydroxyvitamin D3 24-hydroxylase at 6 and 24 h; in contrast 1-hydroxylase increased and was 5 times the control value at 24 h. Finally, the in vitro effects of six cytochrome P-450 inhibitors at concentrations ranging from 10(-7) to 10(-3) M on enzyme activities in renal mitochondrial preparations were compared. Both enzymes were inhibited by all of the inhibitors, but inhibition of 25-hydroxyvitamin D3 24-hydroxylase was consistently greater than that of 25-hydroxyvitamin D3 1-hydroxylase. These studies demonstrate that 24-hydroxylation and 1-hydroxylation respond differently to protein synthesis inhibitors and to cytochrome P-450 inhibitors. The findings are consistent with the hypothesis that the two enzyme activities are associated with different cytochrome P-450 moieties.