Seasonal diet and amphipod size selection of juvenile common bully, <Emphasis Type="Italic">Gobiomorphus cotidianus</Emphasis>, in a coastal New Zealand lake

We examined the seasonal diet of juvenile (≤32 mm total length) common bullies, Gobiomorphus cotidianus, in Lake Waihola, South Island, New Zealand and whether they preyed selectively on the amphipod, Paracalliope fluviatilis, which was a dominant prey item. Generally, ≥75% of all fish sampled had consumed amphipods, which comprised 80%, or more, of the total prey volumes in fish stomachs over most of a year except on one sampling occasion. Copepods, snails, chironomids, Daphnia, mysids and isopods also contributed to the diet. Comparisons of the size distribution of amphipods in fish stomachs and the lake on the date of sampling showed that on four occasions, juvenile bullies selectively consumed small amphipods. No selective feeding was detected on the other occasions. The dominance of primarily benthic amphipods in the diet of juvenile bullies which, in turn, are preyed on by piscivores, suggests a strong benthic-pelagic link in the transfer of energy in Lake Waihola. Handling editor: J. Cambray     

AKT-dependent HspB1 (Hsp27) activity in epidermal differentiation

AKT activity has been reported in the epidermis associated with keratinocyte survival and differentiation. We show in developing skin that Akt activity associates first with post-proliferative, para-basal keratinocytes and later with terminally differentiated keratinocytes that are forming the fetal stratum corneum. In adult epidermis the dominant Akt activity is in these highly differentiated granular keratinocytes, involved in stratum corneum assembly. Stratum corneum is crucial for protective barrier activity, and its formation involves complex and poorly understood processes such as nuclear dissolution, keratin filament aggregation, and assembly of a multiprotein cell cornified envelope. A key protein in these processes is filaggrin. We show that one target of Akt in granular keratinocytes is HspB1 (heat shock protein 27). Loss of epidermal HspB1 caused hyperkeratinization and misprocessing of filaggrin. Akt-mediated HspB1 phosphorylation promotes a transient interaction with filaggrin and intracellular redistribution of HspB1. This is the first demonstration of a specific interaction between HspB1 and a stratum corneum protein and indicates that HspB1 has chaperone activity during stratum corneum formation. This work demonstrates a new role for Akt in epidermis.     

Identification of a redox-sensitive cysteine in GCP60 that regulates its interaction with golgin-160

Golgin-160 is ubiquitously expressed in vertebrates. It localizes to the cytoplasmic side of the Golgi and has a large C-terminal coiled-coil domain. The noncoiled-coil N-terminal head domain contains Golgi targeting information, a cryptic nuclear localization signal, and three caspase cleavage sites. Caspase cleavage of the golgin-160 head domain generates different fragments that can translocate to the nucleus by exposing the nuclear localization signal. We have previously shown that GCP60, a Golgi resident protein, interacts weakly with the golgin-160 head domain but has a strong interaction with one of the caspase-generated golgin-160 fragments (residues 140-311). This preferential interaction increases the Golgi retention of the golgin-160 fragment in cells overexpressing GCP60. Here we studied the interaction of golgin-160-(140-311) with GCP60 and identified a single cysteine residue in GCP60 (Cys-463) that is critical for the interaction of the two proteins. Mutation of the cysteine blocked the interaction in vitro and disrupted the ability to retain the golgin-160 fragment at the Golgi in cells. We also found that Cys-463 is redox-sensitive; in its reduced form, interaction with golgin-160 was diminished or abolished, whereas oxidation of the Cys-463 by hydrogen peroxide restored the interaction. In addition, incubation with a nitric oxide donor promoted this interaction in vitro. These findings suggest that nuclear translocation of golgin-160-(140-311) is a highly coordinated event regulated not only by cleavage of the golgin-160 head but also by the oxidation state of GCP60.     

Escherichia coli O157:H7 colonization at the rectoanal junction of long-duration culture-positive cattle

Long-duration consistently Escherichia coli O157:H7 culture-positive cattle were euthanized and necropsied. Tissue and digesta from along the gastrointestinal tract (GIT) were cultured for the bacteria and examined histologically for lymphoid character. E. coli O157:H7 was detected only at the rectoanal junction mucosa and not at any other GIT location.     

Only one of a wide assortment of manganese-containing SOD mimicking compounds rescues the slow aerobic growth phenotypes of both Escherichia coli and Saccharomyces cerevisiae strains lacking superoxide dismutase enzymes

A variety of manganese-containing coordination compounds, frequently termed superoxide dismutase (SOD) mimics, have been reported to have SOD activity in vitro and to be effective at improving conditions related to increased oxidative stress in multicellular organisms. We tested the effectiveness of several of these compounds in substituting for authentic SOD enzymes in two simple systems--the prokaryote Escherichia coli and the single-celled eukaryote, Saccharomyces cerevisiae--where strains are available that completely lack cytoplasmic SOD activity and are thus significantly impaired in their ability to grow aerobically. Most of the compounds tested, including Euk-8 and Euk-134, manganese salen derivatives developed by Eukarion; M40403, a manganese complex of a bis(cyclohexylpyridine)-substituted macrocyclic ligand developed by Metaphore; and several manganese porphyrin derivatives, were ineffective in both systems. Only the manganese tetrapyridyl porphyrin complex MnTM-2-PyP and two close relatives were effective in rescuing aerobic growth of E. coli lacking SOD, and, in the case of sod1Delta yeast, only MnTM-2-PyP itself was fully effective. Surprisingly, several compounds reported to be beneficial in other in vivo model systems (Euk-8, Euk-134, M40403) were actually toxic to these organisms lacking SOD, although they had no effect on the wild-type parent strains. Our results suggest the possibility that the beneficial effects of some of the so-called "SOD mimic drugs" may be due to some property other than in vivo superoxide dismutase activity.     

Role of renal sympathetic nerve activity in hypertension induced by chronic nitric oxide inhibition

Nitric oxide levels are diminished in hypertensive patients, suggesting nitric oxide might have an important role to play in the development of hypertension. Chronic blockade of nitric oxide leads to hypertension that is sustained throughout the period of the blockade in baroreceptor-intact animals. It has been suggested that the sympathetic nervous system is involved in the chronic increase in blood pressure; however, the evidence is inconclusive. We measured renal sympathetic nerve activity and blood pressure via telemetry in rabbits over 7 days of nitric oxide blockade. Nitric oxide blockade via N(omega)-nitro-L-arginine methyl ester (L-NAME) in the drinking water (50 mg x kg(-1) x day(-1)) for 7 days caused a significant increase in arterial pressure (7 +/- 1 mmHg above control levels; P < 0.05). While the increase in blood pressure was associated with a decrease in heart rate (from 233 +/- 6 beats/min before the L-NAME to 202 +/- 6 beats/min on day 7), there was no change in renal sympathetic nerve activity (94 +/- 4 %baseline levels on day 2 and 96 +/- 5 %baseline levels on day 7 of L-NAME; baseline nerve activity levels were normalized to the maximum 2 s of nerve activity evoked by nasopharyngeal stimulation). The lack of change in renal sympathetic nerve activity during the L-NAME-induced hypertension indicates that the renal nerves do not mediate the increase in blood pressure in conscious rabbits.     

Estradiol and dihydrotestosterone regulate endothelial cell barrier function after hypergravity-induced alterations in MAPK activity

Postflight orthostatic intolerance (POI) was reported to be higher in female than male astronauts and may result from sex-dependent differences in endothelial cell (EC) barrier permeability. Here the effect of 17-estradiol (E₂) and dihydrotestosterone (DHT) on the expression of the tight junction protein occludin, EC barrier function, and MAPK activation over time was tested after subjecting human umbilical vein EC (HUVEC) to brief hypergravity identical to that experienced by astronauts during liftoff (LO) into space. After LO hypergravity, HUVEC showed a time-dependent decrease in occludin correlating with an increase in paracellular permeability and a decrease in transendothelial electrical resistance, indicating a decrease in EC barrier function. LO hypergravity inhibited MAPK activation, which remained suppressed 4 h after LO. Inhibition of MAPK activation correlated with decreased phosphotyrosine occludin, decreased cytochrome-c oxidase activity, and increased paracellular permeability, suggesting a mechanism by which LO hypergravity decreased EC barrier function. Time-dependent differences in MAPK activation, decreased occludin, and EC barrier function between HUVEC treated with E₂ vs. DHT were observed. HUVEC showed delayed activation of MAPK with DHT, i.e., 4 h rather than 2 h for E₂, which correlated with decreased paracellular permeability and the observed sex differences in POI in astronauts. These data temporally separate E₂ and DHT effects in HUVEC and provide evidence for the possible protective roles of sex steroids on EC function after brief exposure to low hypergravity. paracellular permeability; estrogen; androgen     

The apoptosis modulator and tumour suppressor protein RBM5 is a phosphoprotein

RBM5/LUCA-15/H37 is a nuclear SR-related RNA binding protein with the ability to modulate both apoptosis and the cell cycle, and retard tumour formation. How RBM5 functions to carry out these, potentially interrelated, biological activities is unknown. Since reversible phosphorylation has been shown to play an important role in the regulation of SR protein function, apoptosis and cell cycle control, in an attempt to elucidate the underlying mechanisms regulating RBM5 function, the phosphorylation status of RBM5 was investigated. Whole cell lysate from growing cell cultures was treated with the broad phosphatase spectrum of CIP, resulting in a decrease in the molecular mass of RBM5. A similar decrease in molecular mass, of a subset of RBM5 proteins, was observed during growth factor deprivation, in a manner consistent with partial dephosphorylation of RBM5. Molecular mass increased upon growth factor addition, demonstrating that this apoptosis-associated alteration in molecular mass was a reversible process. Immunoprecipitation and mutagenesis experiments strongly suggested that phosphotyrosines are not present in RBM5 under normal growth conditions, and that serine 69 is phosphorylated, but not by Akt kinase. Taken together, these results suggest that reversible phosphorylation of RBM5 is a mechanism capable of regulating RBM5 participation in modulating apoptosis, and perhaps tumour suppression.