Spatial separation of anaerobic ammonium oxidation and nitrite-dependent anaerobic methane oxidation in permeable riverbeds

Anaerobic ammonium oxidation (anammox) and nitrite-dependent anaerobic methane oxidation (n-damo) play important roles in nitrogen and carbon cycling in fresh waters but we do not know how these two processes compete for their common electron acceptor, nitrite. Here, we investigated the spatial distribution of anammox and n-damo across a range of permeable riverbed sediments. Anammox activity and gene abundance were detected in both gravel and sandy riverbeds and showed a simple, common vertical distribution pattern, while the patterns in n-damo were more complex and n-damo activity was confined to the more reduced, sandy riverbeds. Anammox was most active in surficial sediment (0–2 cm), coincident with a peak in hzsA gene abundance and nitrite. In contrast, n-damo activity peaked deeper down (4–8 cm) in the sandy riverbeds, coincident with a peak in n-damo 16S rRNA gene abundance and higher methane concentration. Pore water nitrite, methane and oxygen were key factors influencing the distribution of these two processes in permeable riverbeds. Furthermore, both anammox- and n-damo- activity were positively correlated with denitrification activity, suggesting a role for denitrification in supplying both processes with nitrite. Our data reveal spatial separation between anammox and n-damo in permeable riverbed sediments that potentially avoids them competing for nitrite.     

Cell marking in Arabidopsis thaliana and its application to patch–clamp studies

Ion transport processes at the plasma membrane of plant cells are frequently studied by applying membrane-patch voltage-clamp (patch–clamp) electrophysiological techniques to isolated protoplasts. As plants are composed of many tissues and cell types, and each tissue and cell type may be specialized to a particular function and possess a unique complement of transport proteins, it is important to certify the anatomical origin of the protoplasts used for patch–clamp studies. This paper describes a general molecular genetic approach to marking specific cell types for subsequent patch–clamp studies and presents a specific example: a comparison of the K⁺ currents in protoplasts from cortical and stelar cells of Arabidopsis roots. Transgenic Arabidopsis were generated in which the expression of green fluorescent protein (GFP) from Aequoria victoria was driven by the CaMV 35S promoter (line mGFP3). In roots of the transgenic mGFP3 line, visible fluorescence was restricted to the stele. Protoplasts were generated from roots of the mGFP3 line and K⁺ currents in non-fluorescent (cortical/epidermal) and fluorescent (stelar) protoplasts were assayed using patch–clamp techniques. It was found that both the frequency of observing inward rectifying K⁺ channel (IRC) activity and the relative occurrence of IRC compared to outward rectifying K⁺ channels were significantly lower in protoplasts from cortical/epidermal cells compared to cells of the stele. The presence of GFP did not affect the occurrence or biophysical properties of K⁺ channels. It is concluded that the generation of transgenic Arabidopsis expressing GFP in a cell-specific fashion is a convenient and reliable way to mark protoplasts derived from contrasting cell types for subsequent patch–clamp studies.     

Analysis in a large hyperkalemic periodic paralysis pedigree supports tight linkage to a sodium channel locus.

Hyperkalemic periodic paralysis (HYPP) is an autosomal dominant muscle disease with electrophysiological abnormalities suggesting a defect in a voltage-gated sodium channel (NaCh) gene. A human NaCh gene was recently shown to cosegregate with the disease allele in a family with HYPP. Using an independent clone, we have demonstrated close genetic linkage between an NaCh gene and the HYPP locus in another family. With physiological data demonstrating abnormal NaCh function in HYPP patients, the absence of any obligate recombinations in the two families strengthens the argument that this NaCh gene is the site of the defect in this disorder.     

Estrogen induction of a small, putative K+ channel mRNA in rat uterus

Estrogen causes dramatic long-term changes in the activity of the uterus. Here we report the molecular cloning of a small (700 base) uterine mRNA species capable of inducing a slow K+ current in Xenopus oocytes. The 130 amino acid protein encoded by this mRNA species has a predicted structure that does not resemble that of previously described voltage-dependent channels from mammalian sources. It is, however, similar to structural motifs found in certain prokaryotic ion channels. The induction of this mRNA by estrogen is rapid; this uterine mRNA species is not detectable in uteri from estrogen-deprived rats, but is substantially induced after 3 hr of estrogen treatment. These results support a critical role for regulation of ion channel expression by estrogen in the uterus.     

Negative-Pressure Isolator for Work with Hazardous Infectious Agents in Monkeys

The development, fabrication, and use of a negative-pressure isolator for work with hazardous infectious agents in monkeys or other larger animals are described.