Association of A/T rich microsatellites with responses to artificial selection for larval developmental duration in the silkworm Bombyx mori

Simple sequence repeats (SSRs) and interSSR (ISSR) marker systems were used in this study to reveal genetic changes induced by artificial selection for short/long larval duration in the tropical strain Nistari of the silkworm Bombyx mori. Artificial selection separated longer larval duration (LLD) (29.428 (+/-) 0.723 days) and shorter larval duration (SLD) (22.573 (+/-) 0.839 days) lines from a base, inbred population of Nistari (larval span of 23.143 (+/-) 0.35 days). SSR polymorphism was observed between the LLD and SLD lines at one microsatellite locus, Bmsat106 (CA7) and at two loci of 1074 bp and 823 bp generated with the ISSR primer UBC873. Each of these loci was present only in the LLD line. The loci segregated in the third generation of selection and were fixed in opposite directions. In the F2 generation of the LLD x SLD lines, the alleles of Bmsat106 and UBC8731074bp segregated in a 1:1 ratio and the loci were present only in the LLD individuals. UBC873823bp was homozygous. Single factor ANOVA showed a significant association between the segregating loci and longer larval duration. Together, the two alleles contributed to an 18% increase in larval duration. The nucleotide sequences of the UBC8731074bp and UBC873823bp loci had 67% A/T content and consisted of direct, reverse, complementary and palindromic repeats. The repeats appeared to be "nested" (59%) in larger repeats or as clustered elements adjacent to other repeats. Of 203 microsatellites identified, dinucleotides (67.8%) predominated and were rich in A/T and T/A motifs. The sequences of the UBC8731074bp and UBC873823bp loci showed similarity (E = 0.0) to contigs located in Scaffold 010774 and Scaffold 000139, respectively, of the B. mori genome. BLASTN analysis of the UBC8731074bp sequence showed significant homology of (nt.) 45-122 with upstream region of three exons from Bombyx. The complete sequence of this locus showed approximately 49% nucleotide conservation with transposon 412 of Drosophila melanogaster and the Ikirara insertions of Anopheles gambiae. The A + T richness and lack of coding potential of these small loci, and their absence in the SLD line, reflect the active process of genetic change associated with the switch to short larval duration as an adaptation to the tropics.     

Insights on the role of tillering in salt tolerance of spring wheat from detillering

Tillering is reduced by salinity, with the primary and secondary tillers being more affected than is the mainstem. To understand the importance of tillering in the salt tolerance of wheat plants, two contrasting genotypes of spring wheat (Triticum aestivum L.) were grown in a greenhouse under saline or non-saline conditions and were subjected to five progressive levels of detillering. Regardless of the genotype and salinity, shoot dry weight, seed yield and seed number per plant were all significantly decreased in the treatments where only one or two tillers per plant remained compared with the untouched treatment (more than three tillers), whereas these same variables per tiller tended to be increased on a per tiller (mainstem or substem tiller) basis. The increased seed yield per tiller observed with tiller reduction may be attributed to the enhanced seed number within the spikelet. Under saline conditions, the reductions in shoot dry weight, seed yield and seed number per plant for the salt-tolerant genotype Kharchia were of a greater magnitude in the treatments where only one or two tillers per plant were present compared with the untouched treatment, whereas the magnitude of this reduction in the salt-sensitive genotype Sakha 61 was decreased.     

Spatial and temporal quantitative analysis of cell division and elongation rate in growing wheat leaves under saline conditions

Leaf growth in grasses is determined by the cell division and elongation rates, with the duration of cell elongation being one of the processes that is the most sensitive to salinity. Our objective was to investigate the distribution profiles of cell production, cell length and the duration of cell elongation in the growing zone of the wheat leaf during the steady growth phase. Plants were grown in loamy soil with or without 120 mmol/L NaCl in a growth chamber, and harvested at day 3 after leaf 4 emerged. Results show that the elongation rate of leaf 4 was reduced by 120 mmol/L NaCl during the steady growth phase. The distribution profile of the lengths of abaxial epidermal cells of leaf 4 during the steady growth stage shows a sigmoidal pattern along the leaf axis for both treatments. Although salinity did not affect or even increased the length of the epidermal cells in some locations in the growth zone compared to the control treatment, the final length of the epidermal cells was reduced by 14% at 120 mmol/L NaCl. Thus, we concluded that the observed reduction in the leaf elongation rate derived in part from the reduced cell division rate and either the shortened cell elongation zone or shortened duration of cell elongation. This suggests that more attention should be paid to the effects of salinity on those properties of cell production and the period of cell maturation that are related to the properties of cell wall.     

Polysialic acid regulates the clustering, migration, and neuronal differentiation of progenitor cells in the adult hippocampus

Newborn cells of the adult dentate gyrus in the hippocampus are characterized by their abundant expression of polysialic acid (PSA), a carbohydrate attached to the neural cell adhesion molecule (NCAM). PSA+ newborn cells of the dentate gyrus form clusters with proliferating neural progenitor cells, migrate away from these clusters, and terminally differentiate. To identify the roles of PSA in the development of adult progenitors of the dentate gyrus, we injected endoneuraminidase N (endoN) into the hippocampus of adult rats to specifically cleave PSA from NCAM. Two days later, we administered the mitotic marker, 5-bromo-2'-deoxyuridine (BrdU). Three days after BrdU injection, BrdU+ cells were found inside and outside the clusters of newborn cells. In endoN-treated animals, the total number of BrdU+ cells was not changed but significantly more BrdU+ cells were present within clusters, suggesting that PSA normally facilitates the migration of progenitors away from the clusters. Seven days post-BrdU injection, endoN-treated animals had significantly more BrdU+ cells which were also positive for the mature neuronal nuclear marker NeuN compared with controls, indicating that the loss of PSA from progenitor cells increases neuronal differentiation. This report is the first demonstration that PSA is involved in controlling the spatio-temporal neuronal maturation of adult hippocampal progenitors in the normal brain. In vitro, the removal of PSA from adult-derived neural progenitors significantly enhanced neuronal differentiation, strengthening our in vivo findings and indicating that PSA removal on isolated progenitor cells, apart from a complex in vivo environment, induces neuronal maturation.     

Different mechanisms regulate lysophosphatidic acid (LPA)-dependent versus phorbol ester-dependent internalization of the LPA1 receptor

Lysophosphatidic acid (LPA) stimulates cells by activation of five G-protein-coupled receptors, termed LPA 1-5. The LPA 1 receptor is the most widely expressed and is a major regulator of cell migration. In this study, we show that phorbol ester (PMA)-induced internalization of the LPA(1) receptor requires clathrin AP-2 complexes, protein kinase C, and a distal dileucine motif (amino acids 352 and 353) in the cytoplasmic tail but not beta-arrestin. Agonist-dependent internalization of LPA 1, however, requires a cluster of serine residues (amino acids 341-347) located proximal to the dileucine motif, beta-arrestin, and to a lesser extent clathrin AP-2. The serine cluster of LPA 1 is required for beta-arrestin2-GFP translocation to the plasma membrane and signal desensitization. In contrast, the dileucine motif (IL) is required for both basal and PMA-induced internalization. Evidence for the beta-arrestin independence of PMA-induced internalization of LPA 1 comes from the observations that beta-arrestin2-GFP is not recruited to the plasma membrane upon PMA treatment and that LPA 1 is readily internalized in beta-arrestin1/2 knock-out mouse embryonic fibroblasts. These results indicate that distinct molecular mechanisms regulate agonist-dependent and PMA-dependent internalization of the LPA 1 receptor.     

Molecular Basis of Mucopolysaccharidosis Type IIIB in Emu (Dromaius novaehollandiae): An Avian Model of Sanfilippo Syndrome Type B<

Sanfilippo syndrome type B, or mucopolysaccharidosis (MPS) IIIB, is an autosomal recessive disease caused by a deficiency of lysosomal α-N-acetylglucosaminidase (NAGLU). In Dromaius novaehollandiae (emu), a progressive neurologic disease was recently discovered, which was characterized by NAGLU deficiency and heparan sulfate accumulation. To define the molecular basis, the sequences of the normal emu NAGLU cDNA and gene were determined by PCR-based approaches using primers for highly conserved regions of evolutionarily distant NAGLU homologues. It was observed that the emu NAGLU gene is structurally similar to that of human and mouse, but the introns are considerably shorter. The cDNA had an open reading frame (ORF) of 2259 bp. The deduced amino acid sequence is estimated to share 64% identity with human, 63% with mouse, 41% with Drosophila, 39% with tobacco, and 35% with the Caenorhabditis elegans enzyme. Three normal and two affected emus were studied for nucleotide sequence covering the entire coding region and exon–intron boundaries. Unlike the human gene, emu NAGLU appeared to be highly polymorphic: 19 variations were found in the coding region alone. The two affected emus were found to be homozygous for a 2-bp deletion, 1098-1099delGG, in exon 6. The resulting frameshift predicts a longer ORF of 2370 bp encoding a polypeptide with 37 additional amino acids and 387 altered amino acids. The availability of mutation screening in emus now permits early detection of MPS IIIB in breeding stocks and is an important step in characterizing this unique, naturally occurring avian model for the development of gene transfer studies.     

The role of patient expectations in predicting outcome after total knee arthroplasty

Introduction  

Patient's expectations are variably reported to influence self-rated outcome and satisfaction after medical treatment; this prospective study examined which of the following was the most important unique determinant of global outcome/satisfaction after total knee arthroplasty (TKA): baseline expectations; fulfilment of expectations; or current symptoms and function.     

Quantification of mycorrhizal water uptake via high-resolution on-line water content sensors

The benefits of mycorrhizas for host plants are well known for a large number of species. However, experimental evaluations of the hyphal contribution to the total water uptake and the assessment of the bulk flow velocity in the hyphae are so far contradictory. Barley (Hordeum vulgaris L. Scarlet) with the inoculum Glomus intraradices was grown in a split plant-hyphal chamber with a 5 mm air gap. During the preparation of the chambers with a loamy-silt soil, water content sensors were inserted in each of the plant and the hyphal compartments. These sensors allow non-destructive measurements with high resolution. In total, 8 drying periods with a length of several days were applied with repeated watering following each drying period. A clear decline in water content in the hyphal compartment during each drying period supports the ability of hyphae to transfer water into the plant compartment. The difference between the decline in the hyphal compartment with and without arbuscular mycorrhyzal fungi is significant at the p?<?0.000001 level. The direct and indirect hyphal contribution to the total water uptake was estimated to be about 20%. The application of capacitance sensors for water content determination with a special geometry adapted to the plant-hyphal chambers allows the evaluation of the hyphal water flow with high accuracy.     

Genetic variability and association of ISSR markers with some biochemical traits in mulberry (Morus spp.) genetic resources available in India

Utilizing intersimple sequence repeat (ISSR) markers, 18 mulberry (Morus spp.) germplasm collections were studied for genetic variability, phylogenetic relationship, and association with protein and sugar content. The genetic polymorphism exhibited by ISSR primers was 100%, and the genetic diversity recorded among the mulberry accessions had an average of 0.263 ± 0.094. Dendrogram (unweighted pair group method analysis) clustered the mulberry accessions into two major groups, one comprised the accessions collected from north or northeast regions of India, and the other comprised three subclusters and one isolate, i.e., Assamjati, a collection from Assam. Another subcluster contained accessions collected from Kerala, which belong to Morus indica. These accessions of M. indica from Kerala were found to be genetically diverse from north and northeast India. Multidimensional scaling of the ISSR data clearly separated the mulberry accessions according to their genetic diversity and protein content. Mulberry accessions were arbitrarily grouped into three classes viz. very low, moderate, and high in terms of protein and sugar content using standard statistical programs. Stepwise multiple regression analysis identified four ISSR markers (835_1,600, 835_5,600, 822_2,500, and 807_2,500) associated with protein content with highly positive correlation (p < 0.001) with linear curves with high F values (18.055 to 48.674; p < 0.001). In case of sugar content, four ISSR markers viz. 812₉₀₀, 817_1,500, 826_1,500, and 810_8,000 showed negative correlation. Hence, DNA markers for proteins seem promising and may be used in marker-assisted breeding program.