SMAD versus Non-SMAD Signaling Is Determined by Lateral Mobility of Bone Morphogenetic Protein (BMP) Receptors

Bone (or body) morphogenetic proteins (BMPs) belong to the TGFβ superfamily and are crucial for embryonic patterning and organogenesis as well as for adult tissue homeostasis and repair. Activation of BMP receptors by their ligands leads to induction of several signaling cascades. Using fluorescence recovery after photobleaching, FRET, and single particle tracking microscopy, we demonstrate that BMP receptor type I and II (BMPRI and BMPRII) have distinct lateral mobility properties within the plasma membrane, which is mandatory for their involvement in different signaling pathways. Before ligand binding, BMPRI and a subpopulation of BMPRII exhibit confined motion, reflecting preassembled heteromeric receptor complexes. A second free diffusing BMPRII population only becomes restricted after ligand addition. This paper visualizes time-resolved BMP receptor complex formation and demonstrates that the lateral mobility of BMPRI has a major impact in stabilizing heteromeric BMPRI-BMPRII receptor complexes to differentially stimulate SMAD versus non-SMAD signaling.     

Synthesis of Short-Chain Diols and Unsaturated Alcohols from Secondary Alcohol Substrates by the Rieske Nonheme Mononuclear Iron Oxygenase MdpJ

The Rieske nonheme mononuclear iron oxygenase MdpJ of the fuel oxygenate-degrading bacterial strain Aquincola tertiaricarbonis L108 has been described to attack short-chain tertiary alcohols via hydroxylation and desaturation reactions. Here, we demonstrate that also short-chain secondary alcohols can be transformed by MdpJ. Wild-type cells of strain L108 converted 2-propanol and 2-butanol to 1,2-propanediol and 3-buten-2-ol, respectively, whereas an mdpJ knockout mutant did not show such activity. In addition, wild-type cells converted 3-methyl-2-butanol and 3-pentanol to the corresponding desaturation products 3-methyl-3-buten-2-ol and 1-penten-3-ol, respectively. The enzymatic hydroxylation of 2-propanol resulted in an enantiomeric excess of about 70% for the (R)-enantiomer, indicating that this reaction was favored. Likewise, desaturation of (R)-2-butanol to 3-buten-2-ol was about 2.3-fold faster than conversion of the (S)-enantiomer. The biotechnological potential of MdpJ for the synthesis of enantiopure short-chain alcohols and diols as building block chemicals is discussed.     

Differential regulation by organic compounds and heavy metals of multiple laccase genes in the aquatic hyphomycete Clavariopsis aquatica

To advance the understanding of the molecular mechanisms controlling microbial activities involved in carbon cycling and mitigation of environmental pollution in freshwaters, the influence of heavy metals and natural as well as xenobiotic organic compounds on laccase gene expression was quantified using quantitative real-time PCR (qRT-PCR) in an exclusively aquatic fungus (the aquatic hyphomycete Clavariopsis aquatica) for the first time. Five putative laccase genes (lcc1 to lcc5) identified in C. aquatica were differentially expressed in response to the fungal growth stage and potential laccase inducers, with certain genes being upregulated by, e.g., the lignocellulose breakdown product vanillic acid, the endocrine disruptor technical nonylphenol, manganese, and zinc. lcc4 is inducible by vanillic acid and most likely encodes an extracellular laccase already excreted during the trophophase of the organism, suggesting a function during fungal substrate colonization. Surprisingly, unlike many laccases of terrestrial fungi, none of the C. aquatica laccase genes was found to be upregulated by copper. However, copper strongly increases extracellular laccase activity in C. aquatica, possibly due to stabilization of the copper-containing catalytic center of the enzyme. Copper was found to half-saturate laccase activity already at about 1.8 μM, in favor of a fungal adaptation to low copper concentrations of aquatic habitats.     

HIV and STI Prevalence among Female Sex Workers in C?te d'Ivoire: Why Targeted Prevention Programs Should Be Continued and Strengthened

Objective

To assess condom use and prevalence of STIs and HIV among female sex workers (FSWs), as part of a comprehensive monitoring and evaluation plan of a nationwide sex worker prevention project in Côte d''Ivoire.

Design and Methods

Cross sectional surveys were conducted among FSWs attending five project clinics in Abidjan and San Pedro (2007), and in Yamoussoukro and Gagnoa (2009). A standardized questionnaire was administered in a face-to-face interview, which included questions on socio-demographic characteristics, sexual behaviour and condom use. After the interview, the participants were asked to provide samples for STI and HIV testing.

Results

A total of 1110 FSWs participated in the surveys. There were large differences in socio-demographic and behavioural characteristics between FSW coming for the first time as compared to FSW coming on a routine visit. The prevalence of N. gonorrhoeae or C.trachomatis was 9.1%, 11.8% among first vs. 6.9% routine attendees (p = 0.004). The overall HIV prevalence was 26.6%, it was lower among first time attendees (17.5% as compared to 33.9% for routine attendees, p<0.001). The HIV prevalence among first attendees was also lower than the proportion of HIV positive tests from routine testing and counselling services in the same clinics.

Conclusions

The results show a relatively high STI and HIV prevalence among FSWs in different cities in Côte d''Ivoire. In the light of these results, prevention efforts should continue to focus on FSWs in the country.     

Variable Responses of Lowland Tropical Forest Nutrient Status to Fertilization and Litter Manipulation

Predicting future impacts of anthropogenic change on tropical forests requires a clear understanding of nutrient constraints on productivity. We compared experimental fertilization and litter manipulation treatments in an old-growth lowland tropical forest to distinguish between the effects of inorganic nutrient amendments and changes in nutrient cycling via litterfall. We measured the changes in soil and litter nutrient pools, litterfall, and fine root biomass in plots fertilized with nitrogen (N), phosphorus (P), or potassium (K), and in litter addition and litter removal treatments during 7 years. Soil inorganic N and litter N increased in double-litter plots but not in N-fertilized plots. Conversely, litter P and soil pools of P and K increased in fertilized plots but not in the double-litter plots. Soil and litter pools of N and K decreased in the no-litter plots. Changes in litterfall with added nutrients or litter were only marginally significant, but fine root biomass decreased with both the litter and the K addition. Differences between the two experiments are mostly attributable to the coupled cycling of carbon and nutrients in litter. Increased nutrient inputs in litter may improve plant uptake of some nutrients compared to fertilization with similar amounts. The litter layer also appears to play a key role in nutrient retention. We discuss our findings in the context of possible impacts of anthropogenic change on tropical forests.     

Defining process design space for biotech products: case study of Pichia pastoris fermentation

The concept of "design space" has been proposed in the ICH Q8 guideline and is gaining momentum in its application in the biotech industry. It has been defined as "the multidimensional combination and interaction of input variables (e.g., material attributes) and process parameters that have been demonstrated to provide assurance of quality." This paper presents a stepwise approach for defining process design space for a biologic product. A case study, involving P. pastoris fermentation, is presented to facilitate this. First, risk analysis via Failure Modes and Effects Analysis (FMEA) is performed to identify parameters for process characterization. Second, small-scale models are created and qualified prior to their use in these experimental studies. Third, studies are designed using Design of Experiments (DOE) in order for the data to be amenable for use in defining the process design space. Fourth, the studies are executed and the results analyzed for decisions on the criticality of the parameters as well as on establishing process design space. For the application under consideration, it is shown that the fermentation unit operation is very robust with a wide design space and no critical operating parameters. The approach presented here is not specific to the illustrated case study. It can be extended to other biotech unit operations and processes that can be scaled down and characterized at small scale.     

Fixation procedures for flow cytometric analysis of environmental bacteria

Analysis of environmental bacteria on the single cell level often requires fixation to store the cells and to keep them in a state as near life-like as possible. Fixation procedures should furthermore counteract the increase of autofluorescence, cell clogging, and distortion of surface characteristics. Additionally, they should meet the specific fixation demands of both aerobically and anaerobically grown bacteria. A fixation method was developed based on metal solutions in combination with sodium azide. The fixation efficiencies of aluminium, barium, bismuth, cobalt, molybdenum, nickel, and tungsten salts were evaluated by flow cytometric measurement of the DNA contents as a bacterial population/community stability marker. Statistical equivalence testing was involved to permit highly reliable flow cytometric pattern evaluation. Investigations were carried out with pure cultures representing environmentally important metabolic and respiratory pathways as controls and with activated sludge as an example for highly diverse bacterial communities. A mixture of 5 mM barium chloride and nickel chloride, each and 10% sodium azide was found to be a suitable fixative for all tested bacteria. The described method provided good sample stability for at least 9 days.