Inborn errors of cobalamin metabolism: Effect of cobalamin supplementation in culture on methylmalonyl CoA mutase activity in normal and mutant human fibroblasts

We have examined the effect of addition of hydroxocobalamin to growth medium on the activity of the adenosylcobalamin-requiring enzyme methylmalonyl CoA mutase in normal human fibroblasts and in mutant human fibroblasts derived from patients with inherited methylmalonicacidemia. The mutant cell lines were assigned to four distinct genetic complementation groups (cbl A, cbl B, cbl C, and cbl D), each deficient in some step in the synthesis of adenosylcobalamin from hydroxocobalamin. After control cells were grown in cobalamin-supplemented medium, mutase holoenzyme activity increased markedly in a time- and concentration-dependent fashion. Growth in cobalamin-supplemented medium had no effect on mutase activity in some mutant lines belonging to the cbl B group, while activity increased severalfold in other cbl B mutants and in all cbl A, cbl C, and cbl D mutants examined, although mutase activity was still <10% of control. Comparison of mutase holoenzyme activity and total propionate pathway activity suggests that enhancement of mutase activity in mutant cells after cobalamin supplementation to values 5–10% of control may be sufficient to overcome the inherited metabolic block and to restore total pathway activity to normal.This work was supported in part by a research grant from the National Institutes of Health (AM 12579). H. F. W. is a recipient of a traineeship from the National Institutes of Health (T01-GM02299).     

Rhizosphere soil bacterial communities and nitrogen cycling affected by deciduous and evergreen tree species

Deciduous and evergreen trees differ in their responses to drought and nitrogen (N) demand. Whether or not these functional types affect the role of the bacterial community in the N cycle during drought remains uncertain. Two deciduous tree species (Alnus cremastogyne, an N₂‐fixing species, and Liquidambar formosana) and two evergreen trees (Cunninghamia lanceolata and Pinus massoniana) were used to assess factors in controlling rhizosphere soil bacterial community and N cycling functions. Photosynthetic rates and biomass production of plants, 16S rRNA sequencing and N‐cycling‐related genes of rhizosphere soil were measured. The relative abundance of the phyla Actinobacteria and Firmicutes was higher, and that of Proteobacteria, Acidobacteria, and Gemmatimondaetes was lower in rhizosphere soil of deciduous trees than that of evergreen. Beta‐diversity of bacterial community also significantly differed between the two types of trees. Deciduous trees showed significantly higher net photosynthetic rates and biomass production than evergreen species both at well water condition and short‐term drought. Root biomass was the most important factor in driving soil bacterial community and N‐cycling functions than total biomass and aboveground biomass. Furthermore, 44 bacteria genera with a decreasing response and 46 taxa showed an increased response along the root biomass gradient. Regarding N‐cycle‐related functional genes, copy numbers of ammonia‐oxidizing bacteria (AOB) and autotrophic ammonia‐oxidizing archaea (AOA), N₂ fixation gene (nifH), and denitrification genes (nirK, nirS) were significantly higher in the soil of deciduous trees than in that of the evergreen. Structural equation models explained 50.2%, 47.6%, 48.6%, 49.4%, and 37.3% of the variability in copy numbers of nifH, AOB, AOA, nirK, and nirS, respectively, and revealed that root biomass had significant positive effects on copy numbers of all N‐cycle functional genes. In conclusion, root biomass played key roles in affecting bacterial community structure and soil N cycling. Our findings have important implications for our understanding of plants control over bacterial community and N‐cycling function in artificial forest ecosystems.     

G4-quadruplex-binding proteins: review and insights into selectivity

There are over 700,000 putative G4-quadruplexes (G4Qs) in the human genome, found largely in promoter regions, telomeres, and other regions of high regulation. Growing evidence links their presence to functionality in various cellular processes, where cellular proteins interact with them, either stabilizing and/or anchoring upon them, or unwinding them to allow a process to proceed. Interest in understanding and manipulating the plethora of processes regulated by these G4Qs has spawned a new area of small-molecule binder development, with attempts to mimic and block the associated G4-binding protein (G4BP). Despite the growing interest and focus on these G4Qs, there is limited data (in particular, high-resolution structural information), on the nature of these G4Q-G4BP interactions and what makes a G4BP selective to certain G4Qs, if in fact they are at all. This review summarizes the current literature on G4BPs with regards to their interactions with G4Qs, providing groupings for binding mode, drawing conclusions around commonalities and highlighting information on specific interactions where available.     

Tight junction biogenesis during early development

The tight junction (TJ) is an essential component of the differentiated epithelial cell required for polarised transport and intercellular integrity and signalling. Whilst much can be learnt about how the TJ is constructed and maintained and how it functions using a wide range of cellular systems, the mechanisms of TJ biogenesis within developmental models must be studied to gain insight into this process as an integral part of epithelial differentiation. Here, we review TJ biogenesis in the early mammalian embryo, mainly considering the mouse but also including the human and other species, and, briefly, within the amphibian embryo. We relate TJ biogenesis to inherent mechanisms of cell differentiation and biosynthesis occurring during cleavage of the egg and the formation of the first epithelium. We also evaluate a wide range of exogenous cues, including cell-cell interactions, protein kinase C signalling, gap junctional communication, Na⁺/K⁺-ATPase and cellular energy status, that may contribute to TJ biogenesis in the embryo and how these may shape the pattern of early morphogenesis.     

The mechanistic basis for higher‐order interactions and non‐additivity in competitive communities

Motivated by both analytical tractability and empirical practicality, community ecologists have long treated the species pair as the fundamental unit of study. This notwithstanding, the challenge of understanding more complex systems has repeatedly generated interest in the role of so‐called higher‐order interactions (HOIs) imposed by species beyond the focal pair. Here we argue that HOIs – defined as non‐additive effects of density on per capita growth – are best interpreted as emergent properties of phenomenological models (e.g. Lotka–Volterra competition) rather than as distinct ‘ecological processes’ in their own right. Using simulations of consumer‐resource models, we explore the mechanisms and system properties that give rise to HOIs in observational data. We demonstrate that HOIs emerge under all but the most restrictive of assumptions, and that incorporating non‐additivity into phenomenological models improves the quantitative and qualitative accuracy of model predictions. Notably, we also observe that HOIs derive primarily from mechanisms and system properties that apply equally to single‐species or pairwise systems as they do to more diverse communities. Consequently, there exists a strong mandate for further recognition of non‐additive effects in both theoretical and empirical research.     

CHANGES IN PHOTOSYNTHESIS IN RESPONSE TO COMBINED IRRADIANCE AND TEMPERATURE STRESS IN CYANOBACTERIAL SURFACE WATERBLOOMS1

Buoyant cyanobacteria, previously mixed throughout the water column, float to the lake surface and form a surface waterbloom when mixing subsides. At the surface, the cells are exposed to full sunlight, and this abrupt change in photon irradiance may induce photoinhibition; at the same time, temperature rises as well. This study investigated the damaging effects of this increase in temperature as well as the ecologically more relevant combination of both an increased temperature and a high photon irradiance. Analysis of surface blooms with oxygen microelectrodes showed that integrated oxygen contents that are dependent on the balance of photosynthetic oxygen evolution and respiratory oxygen uptake decreased when temperature was raised above the lake temperature. Gross rates of photosynthesis were unaffected by temperatures up to of 35°C; hence, a moderate increase in temperature mainly stimulated oxygen uptake. Preincubation of cells of the cyanobacterium Anabaena flos-aquae (Lyngb.) de Brébisson at temperatures up to 35°C did not affect the subsequent measurement of rates of net photosynthesis. Another 5°C rise in temperature severely damaged the photosynthetic apparatus. Failure to restore net rates of photosynthesis was coupled to a strong quenching of the ratio of variable to maximum fluorescence, F_v/F_m, that was the result of a rise in F_o. A combination of high temperature and high photon irradiance was more damaging than high temperature alone. In contrast, low photon irradiances offered substantial protection against heat injury of the photosynthetic apparatus. I conclude from this study that because cyanobacteria usually are acclimated to low average irradiance prior to bloom formation, there is a reasonable risk of chronic photoinhibition. The increase in temperature will enhance the photodamage of cells in the top layer of the bloom. Low photon irradiances in subsurface layers will offer protection against heat injury. If the high temperatures extend to the deepest, dark layers of the bloom, damage in those layers is likely to occur.     

Understanding thermostability in cytochrome P450 by combinatorial mutagenesis

The cytochromes P450 are an important class of mono-oxygenases involved in xenobiotic metabolism and steroid biosynthesis in a diverse set of life forms. Discovery of CYP-119, a P450 from the archea Sulfolobus solfataricus has provided a means for understanding nature's method of stabilizing this important protein superfamily. To identify classes of stabilizing interactions used by CYP-119, we have generated a randomized library of point mutants and screened for mutants that are less thermostable than the wild type by monitoring the characteristic Soret band in the visible region of the cell lysis. The selected mutants were characterized by differential scanning calorimetry to compare the temperatures of the melting transitions of the various mutants. The identified mutations suggested that electrostatic interactions involving salt links and charge-charge interactions, as well as contributions from other interactions such as aromatic stacking, and side chain volume of hydrophobic residues contribute to enhanced thermostability in this cytochrome P450.     

Dracula’s ménagerie: A multispecies occupancy analysis of lynx,wildcat, and wolf in the Romanian Carpathians

1. The recovery of terrestrial carnivores in Europe is a conservation success story. Initiatives focused on restoring top predators require information on how resident species may interact with the re‐introduced species as their interactions have the potential to alter food webs, yet such data are scarce for Europe.
2. In this study, we assessed patterns of occupancy and interactions between three carnivore species in the Romanian Carpathians. Romania houses one of the few intact carnivore guilds in Europe, making it an ideal system to assess intraguild interactions and serve as a guide for reintroductions elsewhere.
3. We used camera trap data from two seasons in Transylvanian forests to assess occupancy and co‐occurrence of carnivores using multispecies occupancy models.
4. Mean occupancy in the study area was highest for lynx (Ψ_winter = 0.76 95% CI: 0.42–0.92; Ψ_autumn = 0.71 CI: 0.38–0.84) and wolf (Ψ_winter = 0.60 CI: 0.34–0.78; Ψ_autumn = 0.81 CI: 0.25–0.95) and lowest for wildcat (Ψ_winter = 0.40 CI: 0.19–0.63; Ψ_autumn = 0.52 CI: 0.17–0.78)
5. We found that marginal occupancy predictors for carnivores varied between seasons. We also found differences in predictors of co‐occurrence between seasons for both lynx‐wolf and wildcat‐wolf co‐occurrence. For both seasons, we found that conditional occupancy probabilities of all three species were higher when another species was present.
6. Our results indicate that while there are seasonal differences in predictors of occupancy and co‐occurrence of the three species, co‐occurrence in our study area is high.
7. Terrestrial carnivore recovery efforts are ongoing worldwide. Insights into interspecific relations between carnivore species are critical when considering the depauperate communities they are introduced in. Our work showcases that apex carnivore coexistence is possible, but dependent on protection afforded to forest habitats and their prey base.