Fragmented and scrambled mitochondrial ribosomal RNA coding regions among green algae: a model for their origin and evolution

Mitochondrial ribosomal RNA coding regions in the only three green algal taxa investigated to date are fundamentally different in that they are continuous in Prototheca wickerhamii, but highly fragmented and scrambled in Chlamydomonas reinhardtii and Chlamydomonas eugametos. To gain more insight into the mode of evolution of fragmented and scrambled mitochondrial ribosomal RNA (rRNA) genes within the green algal group, this work (1) provides additional information on fragmentation patterns of mitochondrial small- and large-subunit (SSU and LSU) rRNAs that strongly supports the concept of a gradual increase in the extent of discontinuity of mitochondrial rRNAs among chlorophycean green algae and (2) reports the first example of fragmented and scrambled mitochondrial LSU rRNA coding regions in a green algal taxon outside the Chlamydomonas group. The present study (1) suggests that the scrambling of the mitochondrial rRNA coding regions may have occurred early in the evolution of fragmented and scrambled mitochondrial rRNA genes within the chlorophycean green algal group, most likely in parallel with the fragmentation events, (2) proposes recombination as a possible mechanism involved in the evolution of these mitochondrial rRNA genes, and (3) presents a hypothetical pathway for converting continuous mitochondrial rRNA genes into the highly fragmented and scrambled rRNA coding regions of Chlamydomonas through a series of recombinatorial events between short repeated sequences.      

Histone deacetylase 1 controls cardiomyocyte proliferation during embryonic heart development and cardiac regeneration in zebrafish

In contrast to mammals, the zebrafish maintains its cardiomyocyte proliferation capacity throughout adulthood. However, neither the molecular mechanisms that orchestrate the proliferation of cardiomyocytes during developmental heart growth nor in the context of regeneration in the adult are sufficiently defined yet. We identified in a forward genetic N-ethyl-N-nitrosourea (ENU) mutagenesis screen the recessive, embryonic-lethal zebrafish mutant baldrian (bal), which shows severely impaired developmental heart growth due to diminished cardiomyocyte proliferation. By positional cloning, we identified a missense mutation in the zebrafish histone deacetylase 1 (hdac1) gene leading to severe protein instability and the loss of Hdac1 function in vivo. Hdac1 inhibition significantly reduces cardiomyocyte proliferation, indicating a role of Hdac1 during developmental heart growth in zebrafish. To evaluate whether developmental and regenerative Hdac1-associated mechanisms of cardiomyocyte proliferation are conserved, we analyzed regenerative cardiomyocyte proliferation after Hdac1 inhibition at the wound border zone in cryoinjured adult zebrafish hearts and we found that Hdac1 is also essential to orchestrate regenerative cardiomyocyte proliferation in the adult vertebrate heart. In summary, our findings suggest an important and conserved role of Histone deacetylase 1 (Hdac1) in developmental and adult regenerative cardiomyocyte proliferation in the vertebrate heart.     

Large-scale identification of polymorphic microsatellites using an <Emphasis Type="Italic">in silico</Emphasis> approach

Background  

Simple Sequence Repeat (SSR) or microsatellite markers are valuable for genetic research. Experimental methods to develop SSR markers are laborious, time consuming and expensive. In silico approaches have become a practicable and relatively inexpensive alternative during the last decade, although testing putative SSR markers still is time consuming and expensive. In many species only a relatively small percentage of SSR markers turn out to be polymorphic. This is particularly true for markers derived from expressed sequence tags (ESTs). In EST databases a large redundancy of sequences is present, which may contain information on length-polymorphisms in the SSR they contain, and whether they have been derived from heterozygotes or from different genotypes. Up to now, although a number of programs have been developed to identify SSRs in EST sequences, no software can detect putatively polymorphic SSRs.     

Mental practice-based rehabilitation training to improve arm function and daily activity performance in stroke patients: a randomized clinical trial

Background  

Over 50% of patients with upper limb paresis resulting from stroke face long-term impaired arm function and ensuing disability in daily life. Unfortunately, the number of effective treatments aimed at improving arm function due to stroke is still low. This study aims to evaluate a new therapy for improving arm function in sub-acute stroke patients based on mental practice theories and functional task-oriented training, and to study the predictors for a positive treatment result. It is hypothesized that a six-week, mental practice-based training program (additional to regular therapy) targeting the specific upper extremity skills important to the individual patient will significantly improve both arm function and daily activity performance, as well as being cost effective.     

Role of Maltogenic Amylase and Pullulanase in Maltodextrin and Glycogen Metabolism of Bacillus subtilis 168

The physiological functions of two amylolytic enzymes, a maltogenic amylase (MAase) encoded by yvdF and a debranching enzyme (pullulanase) encoded by amyX, in the carbohydrate metabolism of Bacillus subtilis 168 were investigated using yvdF, amyX, and yvdF amyX mutant strains. An immunolocalization study revealed that YvdF was distributed on both sides of the cytoplasmic membrane and in the periplasm during vegetative growth but in the cytoplasm of prespores. Small carbohydrates such as maltoheptaose and β-cyclodextrin (β-CD) taken up by wild-type B. subtilis cells via two distinct transporters, the Mdx and Cyc ABC transporters, respectively, were hydrolyzed immediately to form smaller or linear maltodextrins. On the other hand, the yvdF mutant exhibited limited degradation of the substrates, indicating that, in the wild type, maltodextrins and β-CD were hydrolyzed by MAase while being taken up by the bacterium. With glycogen and branched β-CDs as substrates, pullulanase showed high-level specificity for the hydrolysis of the outer side chains of glycogen with three to five glucosyl residues. To investigate the roles of MAase and pullulanase in glycogen utilization, the following glycogen-overproducing strains were constructed: a glg mutant with a wild-type background, yvdF glg and amyX glg mutants, and a glg mutant with a double mutant (DM) background. The amyX glg and glg DM strains accumulated significantly larger amounts of glycogen than the glg mutant, while the yvdF glg strain accumulated an intermediate amount. Glycogen samples from the amyX glg and glg DM strains exhibited average molecular masses two and three times larger, respectively, than that of glycogen from the glg mutant. The results suggested that glycogen breakdown may be a sequential process that involves pullulanase and MAase, whereby pullulanase hydrolyzes the α-1,6-glycosidic linkage at the branch point to release a linear maltooligosaccharide that is then hydrolyzed into maltose and maltotriose by MAase.Bacillus subtilis can utilize polysaccharides such as starch, glycogen, and amylose as carbon sources by hydrolyzing them into smaller maltodextrins via the action of extracellular α-amylase (AmyE) (14). In B. subtilis, α-glucosidase encoded by malL has been known to contribute to maltodextrin metabolism in the cell (40, 41). Schönert et al. (42) reported that maltose is transported by the phosphoenolpyruvate-dependent phosphotransferase system (PTS) in B. subtilis. They also reported that maltodextrins with degrees of polymerization (DP) of 3 to 7 (G3 to G7) are taken up via a maltodextrin-specific (Mdx) ATP-binding cassette (ABC) transport system (42). This system is made up of a maltodextrin-binding protein (MdxE) and two membrane proteins (MdxF and MdxG), as well as an ATPase (MsmX). The basic model proposed for the transport and metabolism of maltooligosaccharides includes a series of carbohydrate-hydrolyzing and -transferring enzymes. However, the enzymatic hydrolysis of maltodextrins and glycogen, providing a major energy reservoir in prokaryotes, was not reflected in the model, due probably to a lack of experimental analysis. Unlike those in Bacillus spp., the transport and metabolic systems for maltodextrins in Escherichia coli have been investigated extensively (7, 9, 10). A model for maltose metabolism involving an α-glucanotransferase (MalQ), a maltodextrin glucosidase (MalZ), and a maltodextrin phosphorylase (MalP) was proposed previously based on analyses of the breakdown of ¹⁴C-labeled maltodextrins in various knockout mutants (10).Ninety bacterial genomes were analyzed to identify the enzymes involved in sugar metabolism, and the results suggested that bacterial enzymes for the synthesis and degradation of glycogen belong to the glucosyltransferase and glycosidase/transglycosidase families, respectively. Free-living bacteria such as B. subtilis carry a minimal set of enzymes for glycogen metabolism, encoded by the glg operon of five genes. The four genes most proximal to the promoter encode enzymes for the synthesis of glycogen, including a branching enzyme (glgB), an ADP-glucose phyrophosphorylase (glgC and glcD), and a glycogen synthase (glgA). On the other hand, the most distal gene, glgP, encodes a glycogen phosphorylase (a member of glycosyltransferase family 35) (13, 18), which degrades glycogen branches by forming glucose-1-phosphate (glucose-1-P). B. subtilis carries two additional enzymes encoded at separate loci, a maltogenic amylase (MAase [YvdF, encoded at 304°]) and a pullulanase (AmyX, encoded at 262°), which have been known to degrade glycogen in vitro (15, 31). These two enzymes are ubiquitous among Bacillus spp. and may play an important role in glycogen and maltodextrin metabolism in the bacteria (see Table S1 in the supplemental material).The MAase YvdF in B. subtilis 168 and its homologue in B. subtilis SUH4-2 share 99% identity at both the nucleotide and amino acid sequence levels (4). MAase (EC 3.2.1.133) is a multisubstrate enzyme that acts on substrates such as cyclodextrin (CD), maltooligosaccharides, pullulan, starch, and glycogen (4). MAase belongs to a subfamily of glycoside hydrolase family 13, along with cyclodextrinase (EC 3.2.1.54), neopullulanase (EC 3.2.1.135), and Thermoactinomyces vulgaris R-47 α-amylase II (46). Although the catalytic properties and tertiary structure of MAase have been studied extensively (33), its physiological role in the bacterial cell is yet to be elucidated. The expression pattern of MAase in B. subtilis 168 has been investigated by monitoring the β-galactosidase activity expressed from the yvdF promoter in defined media containing various carbon sources (20). The yvdF promoter is induced in medium containing maltose, starch, or β-CD but is repressed in the presence of glucose, fructose, sucrose, or glycerol as the sole carbon source. In a previous study, Spo0A, a master regulator determining the life cycle of B. subtilis, was shown to be related to the expression of MAase in a positive manner (20). Kiel et al. (18) reported that the glycogen operon in B. subtilis is turned on during sporulation by RNA polymerase containing σ^E. This finding indicated that MAase, along with glycogen phosphorylase and pullulanase, might be involved in the metabolism of maltodextrin and glycogen in vivo.Pullulanases are capable of hydrolyzing the α-1,6-glycosidic linkages of pullulan to form maltotriose (2, 11, 15, 28, 31, 38). In particular, type I pullulanases have been reported to hydrolyze the α-1,6-glycosidic linkages in branched oligosaccharides such as starch, amylopectin, and glycogen, forming maltodextrins linked by α-1,4-glycosidic linkages (11). Pullulanase is also known as a debranching enzyme. The enzymatic properties and three-dimensional structure of AmyX from B. subtilis 168 were investigated by Malle et al. (28). However, to date, the physiological function of pullulanase encoded by amyX has not been investigated.The aim of this study was to elucidate the physiological functions of MAase and pullulanase, specifically concentrating on their roles in the degradation of maltodextrin and glycogen in B. subtilis. For this purpose, studies of the localization of the enzymes, the accumulation of glycogen, and the distribution of glycogen side chains were performed using the wild type and knockouts of MAase- and pullulanase-related genes.     

The genus Teliocrinus (Crinoidea,Echinodermata): a key taxon among pentacrinid stalked crinoids

The main characters of the stalked crinoids of the family Pentacrinitidae attributed to the genus Teliocrinus are re‐evaluated from a quantitative study of phenotype variation, new observations on arm and stalk articulations, and observation of ontogenetic trends. All of the specimens collected in the northern Indian Ocean belong to the same species, i.e. Teliocrinus springeri (Clark, 1909). However, two phenotypes living at different depths remain valid as subspecies: Teliocrinus springeri springeri (Clark, 1909) and Teliocrinus springeri liliaceus (Clark, 1909). Teliocrinus shares several ontogenetic trends with Endoxocrinus, especially in nonfunctional brachial articulations and stalk symplexies. Its assignment to the Diplocrininae is confirmed. A discussion of its affinities with pentacrinid fossil genera in which the crown is well preserved suggests that Diplocrininae could have first appeared during the Lower Cretaceous. A shortening of brachitaxes and a paedomorphic trend of stalk symplexies are the main other evolutionary traits. Nonfunctional articulations are frequently found at the paedomorphic pole of the heterochronic gradient, without clear derived characters. Classification of pentacrinids mainly based on such symplesiomorphy or paedomorphic characters must be definitively abandoned. However, in post‐Palaeozoic stalked crinoids the scarcity of well‐preserved fossils, the high frequency of paedomorphy, and convergent adaptive characters makes phylogenetic reconstruction only based on morphological characters very difficult and speculative. © 2009 The Linnean Society of London, Zoological Journal of the Linnean Society, 2009, 155 , 22–39.     

Comparison of linkage disequilibrium and haplotype diversity on macro- and microchromosomes in chicken

Background

Toll like receptors (TLR) play the central role in the recognition of pathogen associated molecular patterns (PAMPs). Mutations in the TLR1, TLR2 and TLR4 genes may change the ability to recognize PAMPs and cause altered responsiveness to the bacterial pathogens.

Results

The study presents association between TLR gene mutations and increased susceptibility to Mycobacterium avium subsp. paratuberculosis (MAP) infection. Novel mutations in TLR genes (TLR1- Ser150Gly and Val220Met; TLR2 – Phe670Leu) were statistically correlated with the hindrance in recognition of MAP legends. This correlation was confirmed subsequently by measuring the expression levels of cytokines (IL-4, IL-8, IL-10, IL-12 and IFN-γ) in the mutant and wild type moDCs (mocyte derived dendritic cells) after challenge with MAP cell lysate or LPS. Further in silico analysis of the TLR1 and TLR4 ectodomains (ECD) revealed the polymorphic nature of the central ECD and irregularities in the central LRR (leucine rich repeat) motifs.

Conclusion

The most critical positions that may alter the pathogen recognition ability of TLR were: the 9^th amino acid position in LRR motif (TLR1–LRR10) and 4^th residue downstream to LRR domain (exta-LRR region of TLR4). The study describes novel mutations in the TLRs and presents their association with the MAP infection.