Fitness effects of amino acid replacements in the beta-galactosidase of Escherichia coli

Two genetic procedures were used to obtain amino acid replacements in the lacZ-encoded beta-galactosidase in Escherichia coli. Amino acid replacements could be obtained without regard to their effects on lactase activity by selecting spontaneous mutations that relieved the strong polarity of six nonsense mutations. When streaked on MacConkey- lactose indicator plates, approximately 75% of these mutants gave strong red lactose-fermenting colonies, and 25% gave white nonfermenting colonies. Mutants from 11 other nonsense codons were isolated directly using MacConkey-lactose indicator plates, on which positive color indication requires only 0.5% of the wildtype lactase activity. Among the total of 17 codons, 25 variant beta-galactosidases were identified using electrophoresis and thermal denaturation studies. The fitness effects of these variant beta-galactosidases were determined using competition experiments conducted with lactose as the sole nutrient limiting the growth rate in chemostat cultures. Three of the replacements were deleterious, one was selectively advantageous, and the selective effects of the remaining 21 were undetectable under conditions in which the smallest detectable selection coefficient was approximately 0.4%/generation.      

Sweat gland density and response during high-intensity exercise in athletes with spinal cord injuries

Sweat production is crucial for thermoregulation. However, sweating can be problematic for individuals with spinal cord injuries (SCI), as they display a blunting of sudomotor and vasomotor responses below the level of the injury. Sweat gland density and eccrine gland metabolism in SCI are not well understood. Consequently, this study examined sweat lactate (S-LA) (reflective of sweat gland metabolism), active sweat gland density (SGD), and sweat output per gland (S/G) in 7 SCI athletes and 8 able-bodied (AB) controls matched for arm ergometry VO₂peak. A sweat collection device was positioned on the upper scapular and medial calf of each subject just prior to the beginning of the trial, with iodine sweat gland density patches positioned on the upper scapular and medial calf. Participants were tested on a ramp protocol (7 min per stage, 20 W increase per stage) in a common exercise environment (21±1°C, 45-65% relative humidity). An independent t-test revealed lower (p<0.05) SGD (upper scapular) for SCI (22.3 ±14.8 glands · cm⁻²) vs. AB. (41.0 ± 8.1 glands · cm⁻²). However, there was no significant difference for S/G between groups. S-LA was significantly greater (p<0.05) during the second exercise stage for SCI (11.5±10.9 mmol · l⁻¹) vs. AB (26.8±11.07 mmol · l⁻¹). These findings suggest that SCI athletes had less active sweat glands compared to the AB group, but the sweat response was similar (SLA, S/G) between AB and SCI athletes. The results suggest similar interglandular metabolic activity irrespective of overall sweat rate.     

Intraspecific variation influences natural settlement of eastern oysters

As populations decline, their intraspecific diversity also diminishes. Population decline may be exacerbated if a decrease in intraspecific diversity also reduces important ecological functions that maintain population numbers. Oyster reefs are severely overharvested, declining by ~85 % worldwide. We tested how increasing within-species diversity of eastern oysters (Crassostrea virginica) using transplants would affect recruitment of oyster larvae, a key function necessary to maintain future populations. If harvesting reduces population numbers, within-species diversity, and connectivity, then oysters may lose the ability to adapt to changing environmental conditions as well as incur lower levels of recruitment that may hasten their decline. Results from laboratory and field studies indicated that oyster larvae use chemical cues from adult oysters and not from associated fouling communities to select settlement sites. To test how increasing within-species diversity of oysters affected recruitment, we collected oysters from three distinct bay systems in Texas, USA, and compared natural settlement in treatments where all oysters were from a single bay to a mixture of all three bays. Significantly greater recruitment occurred in mixed treatments in 2010, 2011, and 2012 even though oyster recruitment varied by order of magnitude during this time. The net biodiversity effect was positive in all 3 years, indicating that increased recruitment in mixed treatments can be greater than the additive effect of the single bay treatments. Losing intraspecific diversity may reduce recruitment and lead to further declines in oyster populations, illustrating the need for understanding how intraspecific diversity influences ecological functions.     

mTOR Directs Breast Morphogenesis through the PKC-alpha-Rac1 Signaling Axis

Akt phosphorylation is a major driver of cell survival, motility, and proliferation in development and disease, causing increased interest in upstream regulators of Akt like mTOR complex 2 (mTORC2). We used genetic disruption of Rictor to impair mTORC2 activity in mouse mammary epithelia, which decreased Akt phosphorylation, ductal length, secondary branching, cell motility, and cell survival. These effects were recapitulated with a pharmacological dual inhibitor of mTORC1/mTORC2, but not upon genetic disruption of mTORC1 function via Raptor deletion. Surprisingly, Akt re-activation was not sufficient to rescue cell survival or invasion, and modestly increased branching of mTORC2-impaired mammary epithelial cells (MECs) in culture and in vivo. However, another mTORC2 substrate, protein kinase C (PKC)-alpha, fully rescued mTORC2-impaired MEC branching, invasion, and survival, as well as branching morphogenesis in vivo. PKC-alpha-mediated signaling through the small GTPase Rac1 was necessary for mTORC2-dependent mammary epithelial development during puberty, revealing a novel role for Rictor/mTORC2 in MEC survival and motility during branching morphogenesis through a PKC-alpha/Rac1-dependent mechanism.     

Role of the redox state of nicoptinamide adenine dinucleotides in the in vivo regulation of inositol biosynthesis in Neurospora crassa

A possible _vivo role of pyridine nucleotides and their oxidized/reduced ratio on the regulation of inositol biosynthesis in Neurospora crassa was studied.A direct correlation was obtained when the values of of the water-soluble free inositol pool from intact N. crassa mycelia were plotted agains their NAD⁺/NADH or NAD⁺ + NADP⁺/NADH + NADPH ratios. Higher values in this inositol pool coincided with higher values in the chosen ratios.In long-term experiments (48 h), where the mold was grown without shaking, lower values for the inositol pool, the in vitro activity of D-glucose-6-phosphate cycloaldolase (glucocycloaldolase) and the myo-inositol (inositol) in phospholipids were found than those for cells grown with vigorous shaking.In short-time experiements (20 min), using N. crassa cells depleted of endogenous substrates, the in vivo synthesis of inositol was higher in cells incubated with vigorous shaking than in cells incubated without shaking. Nevertheless, in these experiments the in vitro activity of glucocycloaldolase was not affected by the earation conditions.     

Tryptophan excretion by a bradytroph of Hansenula polymorpha growing in methanol.

The growth of a bradytrophic mutant isolated from the methanol-utilizing yeast Hansenula polymorpha was found to be stimulated by both tyrosine and phenylalanine. The mutant contained about half as much chorismate mutase activity as the wild-type strain. When grown in a medium containing 2% mehtanol as the major carbon and energy source, the bradytroph excreted a low but significant level (28 mg/liter) of L-tryptophan.     

Cefonicid as Therapy for Uncomplicated Gonococcal Urethritis Caused by Penicillinase-Producing Neisseria gonorrhoeae

Young men with uncomplicated gonococcal urethritis were treated with 1 gram of cefonicid given intramuscularly plus 1 gram of probenecid by mouth. Of 53 evaluable patients, 33 (62%) had penicillinase-producing Neisseria gonorrhoeae. All but one of these patients were cured. All men who had penicillin-sensitive infections were cured. Cefonicid was highly effective in the treatment of both penicillin-sensitive and penicillin-resistant N gonorrhoeae. Other than moderate pain at the site of injection, there were no adverse side effects. Cefonicid can be added to the group of newer cephalosporins that are effective in the treatment of gonococcal urethritis caused by either penicillin-sensitive or penicillin-resistant strains.