Fine mapping of the MLK-3 gene within 11q13 and its exclusion as the MEN1 susceptibility gene

MLK-3 kinase is a widely expressed serine/ threonine kinase that bears multiple protein interaction domains and regulates signals mediated by the stress-responsive pathway. Thus, MLK-3 signaling affects numerous cellular processes, raising the possibility that MLK-3 might play a role in oncogenesis. In this report, we describe the fine mapping of the MLK-3 gene within the 11q13.1 chromosomal region. By integrating data from somatic cell hybrids and double color fluorescence in situ hybridization on metaphase chromosomes and DNA fibers, MLK-3 has been assigned approximately 1 Mb telomeric of PYGM, close to the D11S546 locus. Since the MEN1 susceptibility locus is also located within the 11q13.1 region, we have carried out Southern and Northern blot analyses, as well as protein truncation assays to establish whether abnormalities in MLK-3 lead to the development of this familial cancer syndrome. Our observations exclude MLK-3 as the MEN1 gene. Received: 25 September 1996 / Revised: 16 December 1996     

Senescence of rice leaves XXXI. Changes of chlorophyll,protein, and polyamine contents and ethylene production during senescence of a chlorophyll-deficient mutant

Leaf senescence of a chlorophylldeficient rice mutant (LT-8) was investigated. At 10 days after planting, the chlorophyll level in the third leaves of rice seedlings of the mutant was about one half that of normal leaves (Norin no. 8), whereas no difference in the protein level could be detected in the two genotypes. The protein level in leaves decreased with increasing age, and no significant difference could be detected during senescence in the two genotypes. Chlorophyll level in the normal leaves also decreased with increasing age. However, the chlorophyll level in the mutant leaves began to decrease only after more than 60% of the initial protein had been degraded. The pattern of ethylene production in the normal leaves was, in general, similar to that in the mutant leaves. Ethylene production first decreased with age, increased to a maximum at day 18, and decreased thereafter. Both spermidine and spermine levels in the leaves of the two genotypes decreased with increasing age. The pattern of the putrescine level in the normal leaves behaved somewhat similar to that in the mutant leaves. However, during the course of senescence, the putrescine level in the mutant leaves was always higher than that in the normal leaves. The possible relationship between endogenous polyamine levels and ethylene production is discussed.     

Melatonin receptor pharmacology: toward subtype specificity

The recent cloning of three distinct melatonin receptor subtypes (Mel_1a, Mel_1b and Mel_1c) which are part of a new family of G-protein coupled receptors, and probably mediate the physiological actions of the hormone, has spurred interest in the design of analogues with subtype selectivity. The 5-methoxyl and N-acetyl groups of melatonin are important for binding to and activation of the receptor. The indole nucleus serves to hold these two groups at the correct distance from one another and allows them to adopt the required orientation for interaction with the receptor binding pocket. We have investigated the subtype selectivity of a number of analogues of melatonin in which the structure has systematically been modified in order to probe the similarities and differences in the interaction of ligand and receptor subtype. At all three subtypes 5-methoxyl and N-acetyl groups of melatonin are important for high affinity binding. However, replacing the 5-methoxyl group (eg with 5-H, 5-OH, 5-Me or 5-BzO) reduces affinity much less at the Mel_1b receptor subtype than at either Mel_1a or Mel_1c cloned subtypes. This suggests differences between the Mel_1b and Mel _1a/1c subtypes in the size and shape of the binding pocket or in the manner in which melatonin interacts with the receptor at this position. Further studies have revealed that analogues with longer N-acyl carbon chains behave similarly at each subtype. These observations suggest that the ‘pocket’ into which the N-acetyl group fits is very similar for each subtype. Substitutions at the 2-position on the indole ring improved affinity at each receptor subtype but did not give selective analogues. The systematic ‘mapping’ of the requirements for binding at each receptor subtype should allow the design of more selective agonists and antagonists, which will be valuable tools for the characterization and classification of functional melatonin receptors.     

Exogenous proline significantly affects the plant growth and nitrogen assimilation enzymes activities in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis>) under salt stress

Salinity has been shown to be a major factor contributing to low nitrogen availability in plants. To verify the changes in nitrogen metabolism activity as affected by the exogenous application of proline under salt stress and its relation to salt tolerance, in vitro rice shoot apices were used as a model to study the growth performance and changes in nitrogen assimilation activities in two Malaysian rice cultivars MR 220 and MR 253. Results revealed that salt stress greatly reduced the plant height, shoot nitrate (NO₃ ^?) content, shoot glutamine synthetase (GS), and root nitrate reductase (NR) activities in both cultivars. Supplementation of proline significantly increased the plant height, number of roots, root NO₃ ^? content, root NR, and root GS activities under salt stress in both cultivars with greater enhancement in MR 253 than MR 220. The results also indicated that MR 253 possessed higher nitrite reductase (NiR) and glutamate synthase (NADH–GOGAT) activities as compared with MR 220 in all tested treatments. It was suggested that the NO₃ ^? content, NR, and GS activities played important roles in regulating nitrogen metabolism under salt stress. Taken together, it was concluded that the ability of proline in mitigating salt stress-induced damages was correlated with the changes in nitrogen assimilation activities.     

Genetic diversity of native and introduced populations of the invasive house crow (<Emphasis Type="Italic">Corvus splendens</Emphasis>) in Asia and Africa

The common house crow (Corvus splendens) is one of the best known and most wide spread species of the family Corvidae. It is a successful invasive species able to exploit urban environments, well removed from its natural distribution. It is considered a pest as it attains high population densities, can cause serious economic losses and has many adverse effects on native fauna and flora, including predation, competitive displacement and disease transmission. Little genetic research on the house crow has been undertaken so we have only a limited understanding of its natural genetic population structure and invasion history. In this study, we employ microsatellite and mitochondrial DNA markers to assess genetic diversity, phylogeography and population structure of C. splendens within its native range represented by Sri Lanka and Bangladesh and introduced range represented by Malaysia, Singapore, Kenya and South Africa. We found high levels of genetic diversity in some of the invasive populations for which multiple invasions are proposed. The lowest genetic diversity was found for the intentionally introduced population in Selangor, Malaysia. Sri Lanka is a possible source population for Malaysia Selangor consistent with a documented introduction over 100 years ago, with port cities within the introduced range revealing possible presence of migrants from other unsampled locations. We demonstrate the power of the approach of using multiple molecular markers to untangle patterns of invasion, provide insights into population structure and phylogeographic relationships and illustrate how historical processes may have contributed to making this species such a successful invader.     

Requirement for Phosphoglucose Isomerase of Xanthomonas campestris in Pathogenesis of Citrus Canker

A mutant (XT906) of Xanthomonas campestris pv. citri, the causal agent of citrus canker, was induced by insertion of the transposon Tn5tac1 and isolated. This mutant did not grow or elicit canker disease in citrus leaves but was still able to induce a hypersensitive response in a nonhost plant (the common bean). The mutant was also unable to grow on minimal medium containing fructose or glycerol as the sole carbon source. A 2.5-kb fragment of wild-type DNA that complemented the mutant phenotype of XT906 was isolated. Sequence analysis revealed that this DNA fragment encoded a protein of 562 amino acids that shows homology to phosphoglucose isomerase (PGI). Enzyme activity assay confirmed that the encoded protein possesses PGI activity. Analysis of the activity of the promoter of the pgi gene revealed that it was inhibited by growth in complex medium but induced by culture in plant extract. These results demonstrate that PGI is required for pathogenicity of X. campestris pv. citri.     

Novel strain-level resolution of Crohn’s disease mucosa-associated microbiota via an ex vivo combination of microbe culture and metagenomic sequencing

The mucosa-associated microbiota is widely recognized as a potential trigger for Crohn’s disease pathophysiology but remains largely uncharacterised beyond its taxonomic composition. Unlike stool microbiota, the functional characterisation of these communities using current DNA/RNA sequencing approaches remains constrained by the relatively small microbial density on tissue, and the overwhelming amount of human DNA recovered during sample preparation. Here, we have used a novel ex vivo approach that combines microbe culture from anaerobically preserved tissue with metagenome sequencing (MC-MGS) to reveal patient-specific and strain-level differences among these communities in post-operative Crohn’s disease patients. The 16 S rRNA gene amplicon profiles showed these cultures provide a representative and holistic representation of the mucosa-associated microbiota, and MC-MGS produced both high quality metagenome-assembled genomes of recovered novel bacterial lineages. The MC-MGS approach also produced a strain-level resolution of key Enterobacteriacea and their associated virulence factors and revealed that urease activity underpins a key and diverse metabolic guild in these communities, which was confirmed by culture-based studies with axenic cultures. Collectively, these findings using MC-MGS show that the Crohn’s disease mucosa-associated microbiota possesses taxonomic and functional attributes that are highly individualistic, borne at least in part by novel bacterial lineages not readily isolated or characterised from stool samples using current sequencing approaches.Subject terms: Biomarkers, Microbiome, Next-generation sequencing, Clinical microbiology, Metagenomics