Simultaneous increase in cellular content and volumetric concentration of lipids in <Emphasis Type="Italic">Bracteacoccus bullatus</Emphasis> cultivated at reduced nitrogen and phosphorus concentrations

Manipulation of the nutrient concentration is an inexpensive and efficient method for increasing lipid and TAG accumulation in algal cells. However, high volumetric production requires finding a proper balance between the decrease of biomass production and the increase in the total lipid content. We isolated a strain of green microalga Bracteacoccus bullatus and increased its lipid content from 17 to 59% of biomass dry weight by manipulating of nitrogen and phosphorus content in the medium. The 10-fold reduction of the nitrogen and phosphorus concentration in the medium was the most efficient method of the lipid induction compared to nutrient deplete and high nutrient conditions. The oleic (48–64% mass of total fatty acids) and linoleic (14–24% mass of total fatty acids) acids dominated in the fatty acid profile, thus making this strain a suitable candidate for biodiesel production.     

Persistence of social jetlag and sleep disruption in healthy young adults

Sleep disruption has been associated with increased risks for several major chronic diseases that develop over decades. Differences in sleep/wake timing between work and free days can result in the development of social jetlag (SJL), a chronic misalignment between a person’s preferred sleep/wake schedule and sleep/wake timing imposed by his/her work schedule. Only a few studies have examined the persistence of SJL or sleep disruption over time. This prospective investigation examined SJL and sleep characteristics over a 2-year period to evaluate whether SJL or poor sleep were chronic conditions during the study period. SJL and sleep measures (total sleep time [TST], sleep onset latency [SOL], wake after sleep onset [WASO]), and sleep efficiency [SE]), were derived from armband monitoring among 390 healthy men and women 21–35 years old. Participants wore the armband for periods of 4–10 days at 6-month intervals during the follow-up period (N = 1431 repeated observations).

The consistency of SJL or sleep disruption over time was analyzed using generalized linear mixed models (GLMMs) for repeated measures. Repeated measures latent class analysis (RMLCA) was then used to identify subgroups among the study participants with different sleep trajectories over time. Individuals in each latent group were compared using GLMMs to identify personal characteristics that differed among the latent groups.

Minor changes in mean SJL, chronotype, or TST were observed over time, whereas no statistically significant changes in SOL, WASO, or SE were observed during the study period. The RMLCA identified two groups of SJL that remained consistent throughout the study (low SJL, mean ± SE: 0.4 ± 0.04 h, 42% of the study population; and high SJL, 1.4 ± 0.03 h, 58%). Those in the SJL group with higher values tended to be employed and have an evening chronotype.

Similarly, two distinct subgroups were observed for SOL, WASO, and SE; one group with a pattern suggesting disrupted sleep over time, and another with a consistently normal sleep pattern. Analyses of TST identified three latent groups with relatively short (5.6 ± 1.0 h, 21%), intermediate (6.5 ± 1.0 h, 44%), and long (7.3 ± 1.0 h, 36%) sleep durations, all with temporally stable, linear trajectories. The results from this study suggest that sleep disturbances among young adults can persist over a 2 year period. Latent groups with poor sleep tended to be male, African American, lower income, and have an evening chronotype relative to those with more normal sleep characteristics. Characterizing the persistence of sleep disruption over time and its contributing factors could be important for understanding the role of poor sleep as a chronic disease risk factor.     

Resistance training increases basal limb blood flow and vascular conductance in aging humans.

Age-related reductions in basal limb blood flow and vascular conductance are associated with the metabolic syndrome, functional impairments, and osteoporosis. We tested the hypothesis that a strength training program would increase basal femoral blood flow in aging adults. Twenty-six sedentary but healthy middle-aged and older subjects were randomly assigned to either a whole body strength training intervention group (52 +/- 2 yr, 3 men, 10 women) who underwent three supervised resistance training sessions per week for 13 wk or a control group (53 +/- 2 yr, 4 men, 9 women) who participated in a supervised stretching program. At baseline, there were no significant differences in blood pressure, cardiac output, basal femoral blood flow (via Doppler ultrasound), vascular conductance, and vascular resistance between the two groups. The strength training group increased maximal strength in all the major muscle groups tested (P < 0.05). Whole body lean body mass increased (P < 0.05) with strength training, but leg fat-free mass did not. Basal femoral blood flow and vascular conductance increased by 55-60% after strength training (both P < 0.05). No such changes were observed in the control group. In both groups, there were no significant changes in brachial blood pressure, plasma endothelin-1 and angiotensin II concentrations, femoral artery wall thickness, cardiac output, and systemic vascular resistance. Our results indicate that short-term strength training increases basal femoral blood flow and vascular conductance in healthy middle-aged and older adults.     

An efficient survey method for estimating populations of Marsh Tits Poecile palustris,a low-density woodland passerine

Capsule: A playback survey comprising two visits to woodland in early spring can reliably detect Marsh Tits Poecile palustris and permit reasonable estimation of the number of territories.

Aims: To assess the efficacy of an efficient survey method for detecting and estimating populations of Marsh Tits.

Methods: Detection probability of colour-marked Marsh Tits, surveyed using playback, was assessed with Cormack–Jolly–Seber models. Reliability of territory estimates was compared between colour-marked and largely or wholly unmarked populations.

Results: Playback surveys over two visits in early spring were highly effective in detecting individual Marsh Tits. Territory estimates were similar for woods in years where less than 20% of the population was marked compared to years when a higher proportion of birds were colour-marked, although territories may be underestimated in larger woods with unmarked populations.

Conclusion: A playback survey comprising two visits in early spring is recommended as an efficient method of surveying Marsh Tit populations and locating individuals. A survey protocol is suggested.     

Mechanism of interactions of alpha-naphthoflavone with cytochrome P450 3A4 explored with an engineered enzyme bearing a fluorescent probe

Design of a partially cysteine-depleted C98S/C239S/C377S/C468A cytochrome P450 3A4 mutant designated CYP3A4(C58,C64) allowed site-directed incorporation of thiol-reactive fluorescent probes into alpha-helix A. The site of modification was identified as Cys-64 with the help of CYP3A4(C58) and CYP3A4(C64), each bearing only one accessible cysteine. Changes in the fluorescence of CYP3A4(C58,C64) labeled with 6-(bromoacetyl)-2-(dimethylamino)naphthalene (BADAN), 7-(diethylamino)-3-(4'-maleimidylphenyl)-4-methylcoumarin (CPM), or monobromobimane (mBBr) were used to study the interactions with bromocriptine (BCT), 1-pyrenebutanol (1-PB), testosterone (TST), and alpha-naphthoflavone (ANF). Of these substrates only ANF has a specific effect, causing a considerable decrease in fluorescence intensity of BADAN and CPM and increasing the fluorescence of mBBr. This ANF-binding event in the case of the BADAN-modified enzyme is characterized by an S50 of 18.2 +/- 0.7, compared with the value of 2.2 +/- 0.3 for the ANF-induced spin transition, thus revealing an additional low-affinity binding site. Studies of the effect of TST, 1-PB, and BCT on the interactions of ANF monitored by changes in fluorescence of CYP3A4(C58,C64)-BADAN or by the ANF-induced spin transition revealed no competition by these substrates. Investigation of the kinetics of fluorescence increase upon H2O2-dependent heme depletion suggests that labeled CYP3A4(C58,C64) is represented by two conformers, one of which has the fluorescence of the BADAN and CPM labels completely quenched, presumably by photoinduced electron transfer from the neighboring Trp-72 and/or Tyr-68 residues. The binding of ANF to the newly discovered binding site appears to affect the interactions of the label with the above residue(s), thus modulating the fraction of the fluorescent conformer.     

Reflux of Endoplasmic Reticulum proteins to the cytosol inactivates tumor suppressors

In the past decades, many studies reported the presence of endoplasmic reticulum (ER)‐resident proteins in the cytosol. However, the mechanisms by which these proteins relocate and whether they exert cytosolic functions remain unknown. We find that a subset of ER luminal proteins accumulates in the cytosol of glioblastoma cells isolated from mouse and human tumors. In cultured cells, ER protein reflux to the cytosol occurs upon ER proteostasis perturbation. Using the ER luminal protein anterior gradient 2 (AGR2) as a proof of concept, we tested whether the refluxed proteins gain new functions in the cytosol. We find that refluxed, cytosolic AGR2 binds and inhibits the tumor suppressor p53. These data suggest that ER reflux constitutes an ER surveillance mechanism to relieve the ER from its contents upon stress, providing a selective advantage to tumor cells through gain‐of‐cytosolic functions—a phenomenon we name ER to Cytosol Signaling (ERCYS).     

The betagamma subunit of heterotrimeric G proteins interacts with RACK1 and two other WD repeat proteins

A yeast two-hybrid approach was used to discern possible new effectors for the betagamma subunit of heterotrimeric G proteins. Three of the clones isolated are structurally similar to Gbeta, each exhibiting the WD40 repeat motif. Two of these proteins, the receptor for activated C kinase 1 (RACK1) and the dynein intermediate chain, co-immunoprecipitate with Gbetagamma using an anti-Gbeta antibody. The third protein, AAH20044, has no known function; however, sequence analysis indicates that it is a WD40 repeat protein. Further investigation with RACK1 shows that it not only interacts with Gbeta(1)gamma(1) but also unexpectedly with the transducin heterotrimer Galpha(t)beta(1)gamma(1). Galpha(t) alone does not interact, but it must contribute to the interaction because the apparent EC(50) value of RACK1 for Galpha(t)beta(1)gamma(1) is 3-fold greater than that for Gbeta(1)gamma(1) (0.1 versus 0.3 microm). RACK1 is a scaffold that interacts with several proteins, among which are activated betaIIPKC and dynamin-1 (1). betaIIPKC and dynamin-1 compete with Gbeta(1)gamma(1) and Galpha(t)beta(1)gamma(1) for interaction with RACK1. These findings have several implications: 1) that WD40 repeat proteins may interact with each other; 2) that Gbetagamma interacts differently with RACK1 than with its other known effectors; and/or 3) that the G protein-RACK1 complex may constitute a signaling scaffold important for intracellular responses.