Flavonoids: a colorful model for the regulation and evolution of biochemical pathways

For more than a century, the biosynthesis of flavonoid pigments has been a favorite of scientists to study a wide variety of biological processes, such as inheritance and transposition, and has become one of the best-studied pathways in nature. The analysis of pigmentation continues to provide insights into new areas, such as the channeling and intracellular transport of metabolites, regulation of gene expression and RNA interference. Moreover, because pigmentation is studied in a variety of species, it provides unique molecular insights into the evolution of biochemical pathways and regulatory networks.     

Identification of Enterococcus spp. with a biochemical key.

A six-step biochemical key is presented for the identification of all recognized Enterococcus spp. The key consists of 12 tests, but no more than 6 are needed for the most complicated identification. The reliability of the key has been evaluated with collection type strains and clinical and environmental isolates. This key has fewer tests than those reported in previous studies. There is no commercial kit that includes the whole set of tests. However, some of the tests are included in enzyme activity-based kits that could be used with the proposed key. The key is designed for use in routine applications, especially in environmental and clinical studies with a high number of isolates.     

Embolism resistance as a key mechanism to understand adaptive plant strategies

Fine-scale interconduit pit modifications regulating drought-induced embolism.

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Highlights► The hydraulic pathway of plants is vulnerable to develop air bubbles. ► Air bubble formation in plants is caused by drought or freeze–thaw events. ► Various mechanical xylem properties are correlated with drought-induced embolisms. ► Angiosperms have a greater ability to repair stem embolisms than gymnosperms. ► Secondarily woody shrubs are more embolism resistant than herbaceous relatives.     

Signals, pathways and splicing regulation

Alternative splicing of messenger RNA precursors is an extraordinary source of protein diversity and the regulation of this process is crucial for diverse cellular functions in both physiological and pathological situations. For many years, several signaling pathways have been implicated in alternative splicing regulation. Recent work has begun to unravel the molecular mechanisms by which extracellular stimuli activate signaling cascades that modulate the activity of the splicing machinery and therefore the splicing pattern of many different target messenger RNA precursors. These experiments are revealing unexpected aspects of the mechanism that control splicing and the consequences of the regulated splicing events. We summarize here the current knowledge about signal-induced alternative splicing regulation of Slo, NR1, CD44, CD45 and fibronectin genes, and also discuss the importance of some of these events in determination of cellular fate. Furthermore, we highlight the relevance of signal-induced changes in phosphorylation state and subcellular distribution of splicing factors as a way of regulating the splicing process. Lastly, we explore new and unexpected findings about regulated splicing in anucleated cells.     

The NP-C gene: a key to pathways of intracellular cholesterol transport

Elucidation of the pathways for intracellular transport of cholesterol is an important yet elusive goal in cell biology. Analysis of the cellular defects in the human disease Niemann-Pick C (NP-C) is providing insights into this problem. Cholesterol derived from low-density lipoprotein accumulates in lysosomes of NP-C cells, apparently because intracellular movement of such cholesterol is blocked. Identification of the NP-C gene should provide crucial molecular clues to the mechanism of cholesterol transport within cells.     

Integrative analysis of transcript and metabolite profiling data sets to evaluate the regulation of biochemical pathways during photomorphogenesis

One of the key developmental processes during photomorphogenesis is the differentiation of prolamellar bodies of proplastids into thylakoid membranes containing the photosynthetic pigment-protein complexes of chloroplasts. To study the regulatory events controlling pigment-protein complex assembly, including the biosynthesis of metabolic precursors and pigment end products, etiolated Arabidopsis thaliana seedlings were irradiated with continuous red light (Rc), which led to rapid greening, or continuous far-red light (FRc), which did not result in visible greening, and subjected to analysis by oligonucleotide microarrays and targeted metabolite profiling. An analysis using BioPathAt, a bioinformatic tool that allows the visualization of post-genomic data sets directly on biochemical pathway maps, indicated that in Rc-treated seedlings mRNA expression and metabolite patterns were tightly correlated (e.g., Calvin cycle, biosynthesis of chlorophylls, carotenoids, isoprenoid quinones, thylakoid lipids, sterols, and amino acids). K-means clustering revealed that gene expression patterns across various biochemical pathways were very similar in Rc- and FRc-treated seedlings (despite the visible phenotypic differences), whereas a principal component analysis of metabolite pools allowed a clear distinction between both treatments (in accordance with the visible phenotype). Our results illustrate the general importance of integrative approaches to correlate post-genomic data sets with phenotypic outcomes.     

Allosteric regulation, cooperativity, and biochemical oscillations

Allosteric regulation is associated with a number of periodic phenomena in biochemical systems. The cooperative nature of such regulatory interactions provides a source of nonlinearity that favors oscillatory behavior. We assess the role of cooperativity in the onset of biochemical oscillations by analyzing two specific examples. First, we consider a model for a product-activated allosteric enzyme which has previously been proposed to account for glycolytic oscillations. While enzyme cooperativity plays an important role in the occurrence of oscillations, we show that these may nevertheless occur in the absence of cooperativity when the reaction product is removed in a Michaelian rather than linear manner. The second model considered was recently proposed to account for signal-induced oscillations of intracellular calcium. This phenomenon originates from a nonlinear process of calcium-induced calcium release. Here also, the cooperative nature of that positive feedback favors the occurrence of oscillations but is not absolutely required for periodic behavior. Besides underlining the importance of cooperativity, the results highlight the role of diffuse nonlinearities distributed over several steps within a regulated system: even in the absence of cooperativity, such mild nonlinearities (e.g., of the Michaelian type) may combine to raise the overall degree of nonlinearity up to the level required for oscillations.     

Biological thiols elicit prolactin proteolysis by glandular kallikrein and permit regulation by biochemical pathways linked to redox control

Rat glandular kallikrein (GK), a trypsin-like serine protease, cleaves rat prolactin (PRL) in vitro to novel forms detectable in vivo and likely to be of physiological significance. PRL proteolysis by GK is thiol-dependent, with thiols acting upon PRL to refold the molecule into novel conformations that are GK substrates. This study compared several natural and synthetic thiols for their ability to elicit PRL proteolysis by GK. Rat PRL was incubated with rat GK in the presence of various thiols and 0.5% Triton X-100, which enhances thiol-elicited proteolysis. Cleavage was analyzed by gel electrophoresis under reducing and nonreducing conditions. In the presence of Triton X-100, all low molecular weight thiols elicited PRL cleavage by GK. The order of potency was dithiothreitol greater than mercaptoethanol greater than lipoic acid greater than cysteamine = glutathione (GSH) = coenzyme A greater than cysteine. In the absence of Triton, however, dithiothreitol, coenzyme A, and mercaptoethanol were most effective in eliciting GK proteolysis. Triton X-100 enhanced PRL cleavage by 4-19-fold, depending upon the thiol used. Folding isomers of processed PRL observed following cleavage included disulfide-liked homodimers, oxidized monomers, reduced monomers and mixed disulfides; the folding isomers generated varied depending upon the thiol used. GSH potency in eliciting PRL proteolysis increased 10-fold in the presence of biochemical pathways shuttling reducing equivalents to GSH disulfide (GSSG). PRL cleavage by GK could be controlled by substrates, enzymes, and cofactors making up the reducing shuttle when GSSG was used. Thioredoxin (a protein disulfide oxidoreductase) potently elicited PRL proteolysis by GK in the presence of a reducing shuttle and Triton X-100.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cellular models to investigate biochemical pathways in Parkinson's disease

Cellular models are instrumental in dissecting a complex pathological process into simpler molecular events. Parkinson's disease is multifactorial and clinically heterogeneous; the aetiology of the sporadic (and most common) form is still unclear and only a few molecular mechanisms have been clarified so far in the neurodegenerative cascade. In such a multifaceted picture, it is particularly important to identify experimental models that simplify the study of the different networks of proteins/genes involved. Cellular models that reproduce some of the features of the neurons that degenerate in Parkinson's disease have contributed to many advances in our comprehension of the pathogenic flow of the disease. In particular, the pivotal biochemical pathways (i.e. apoptosis and oxidative stress, mitochondrial impairment and dysfunctional mitophagy, unfolded protein stress and improper removal of misfolded proteins) have been widely explored in cell lines, challenged with toxic insults or genetically modified. The central role of α-synuclein has generated many models aiming to elucidate its contribution to the dysregulation of various cellular processes. In conclusion, classical cellular models appear to be the correct choice for preliminary studies on the molecular action of new drugs or potential toxins and for understanding the role of single genetic factors. Moreover, the availability of novel cellular systems, such as cybrids or induced pluripotent stem cells, offers the chance to exploit the advantages of an in vitro investigation, although mirroring more closely the cell population being affected.     

Training-induced adaptive plasticity in human somatosensory reflex pathways.

This paper reviews evidence supporting adaptive plasticity in muscle and cutaneous afferent reflex pathways induced by training and rehabilitative interventions. The perspective is advanced that the behavioral and functional relevance of any intervention and the reflex pathway under study should be considered when evaluating both adaptation and transfer. A cornerstone of this concept can be found in acute task-dependent reflex modulation. Because the nervous system allows the expression of a given reflex according to the motor task, an attempt to evaluate the training adaptation should also be evoked under the same conditions as training bearing in mind the functional role of the pathway under study. Within this framework, considerable evidence supports extensive adaptive plasticity in human muscle afferent pathways in the form of operant conditioning, strength training, skill training, and locomotor training or retraining. Directly comparable evidence for chronic adaptation in cutaneous reflex pathways is lacking. However, activity-dependent plasticity in cutaneous pathways is documented particularly in approaches to neurological rehabilitation. Overall, the adaptive range for human muscle afferent reflexes appears bidirectional (that is, increased or reduced amplitudes) and on the order of 25-50%. The adaptive range for cutaneous pathways is currently uncertain.