Deafness-related plasticity in the inferior colliculus: gene expression profiling following removal of peripheral activity

The inferior colliculus (IC) is a major center of integration in the ascending as well as descending auditory pathways, where both excitatory and inhibitory amino acid neurotransmitters play a key role. When normal input to the auditory system is decreased, the balance between excitation and inhibition in the IC is disturbed. We examined global changes in gene expression in the rat IC 3 and 21 days following bilateral deafening, using Affymetrix GeneChip arrays and focused our analysis on changes in expression of neurotransmission-related genes. Over 1400 probe sets in the Affymetrix Rat Genome U34A Array were identified as genes that were differentially expressed. These genes encoded proteins previously reported to change as a consequence of deafness, such as calbindin, as well as proteins not previously reported to be modulated by deafness, such as clathrin. A subset of 19 differentially expressed genes was further examined using quantitative RT-PCR at 3, 21 and 90 days following deafness. These included several GABA, glycine, glutamate receptor and neuropeptide-related genes. Expression of genes for GABA-A receptor subunits beta2, beta3, and gamma2, plus ionotropic glutamate receptor subunits AMPA 2, AMPA 3, and kainate 2, increased at all three times. Expression of glycine receptor alpha1 initially declined and then later increased, while alpha2 increased sharply at 21 days. Glycine receptor alpha3 increased between 3 and 21 days, but decreased at 90 days. Of the neuropeptide-related genes tested with qRT-PCR, tyrosine hydroxylase decreased approximately 50% at all times tested. Serotonin receptor 2C increased at 3, 21, and 90 days. The 5B serotonin receptor decreased at 3 and 21 days and returned to normal by 90 days. Of the genes tested with qRT-PCR, only glycine receptor alpha2 and serotonin receptor 5B returned to normal levels of expression at 90 days. Changes in GABA receptor beta3, GABA receptor gamma2, glutamate receptor 2/3, enkephalin, and tyrosine hydroxylase were further confirmed using immunocytochemistry.     

ANG II type 2 receptor regulates smooth muscle growth and force generation in late fetal mouse development

Although evidence from culture studies implicates the angiotensin II (ANG II) type 2 receptor (AT(2)R) in the regulation of growth and differentiation of arterial smooth muscle (SM) cells (SMC), the lack of its expression in adult arteries has precluded direct investigation of its role in vivo. The goal of the present study was to determine the role of AT(2)R in the control of fetal SMC growth, contractility, and differentiation during vascular development. Determination of isometric tension in fetal aortas showed potentiated ANG II-induced contraction by treatment with the selective AT(2)R antagonist PD-123319, demonstrating the presence of functional AT(2)Rs that mediate reduced force development in vascular SMC. In direct contrast to numerous cell culture studies, proliferation indexes were decreased rather than increased in aortic SMC of fetal homozygous AT(2)R knockout compared with wild-type or heterozygous knockout mice. Experiments using SMC tissues from heterozygous female AT(2)R knockout mice, which are naturally occurring chimeras for AT(2)R expression, showed that AT(2)R mRNA expression was exactly 50% of that of wild type. This indicated that loss of AT(2)R expression did not confer a selective advantage or disadvantage for SMC lineage determination and expansion. Real time RT-PCR analyses showed no significant difference in expression of SM-alpha-actin, SM myosin heavy chain, and myocardin in various SM tissues from all three genotypes, suggesting that knockout of AT(2)R had no effect on subsequent SMC differentiation. Taken together, results indicate that functional AT(2)R are expressed in fetal aorta and mediate reduced force development but do not significantly contribute to regulation of SMC differentiation.     

Chromosome evolution in eukaryotes: a multi-kingdom perspective

In eukaryotes, chromosomal rearrangements, such as inversions, translocations and duplications, are common and range from part of a gene to hundreds of genes. Lineage-specific patterns are also seen: translocations are rare in dipteran flies, and angiosperm genomes seem prone to polyploidization. In most eukaryotes, there is a strong association between rearrangement breakpoints and repeat sequences. Current data suggest that some repeats promoted rearrangements via non-allelic homologous recombination, for others the association might not be causal but reflects the instability of particular genomic regions. Rearrangement polymorphisms in eukaryotes are correlated with phenotypic differences, so are thought to confer varying fitness in different habitats. Some seem to be under positive selection because they either trap favorable allele combinations together or alter the expression of nearby genes. There is little evidence that chromosomal rearrangements cause speciation, but they probably intensify reproductive isolation between species that have formed by another route.     

Light in retinitis pigmentosa

Retinitis pigmentosa (RP) is one of the most genetically heterogeneous inherited disorders. Twelve genes have now been identified in the autosomal dominant form of the disease, including some recently characterized genes that show unprecedented and fascinating traits in both their function and in their expression profiles. These include many widely expressed genes encoding components of the spliceosome and a guanine nucleotide synthesis gene. Intriguingly, the most recently identified dominant gene does not appear to be expressed in the neuronal retina but is expressed in the capillaries of the choroid. In attempting to understand the effects of mutations in these genes, investigators are forced to re-evaluate their thinking on the molecular mechanisms of genetic blindness and to undertake an increasingly inter-disciplinary approach in their analysis of this disease. Recently, this has resulted in significant developments in the elucidation of the molecular pathogenesis of RP.     

PCR-based microsatellite analysis for differentiation and genetic monitoring of nine inbred SENCAR mouse strains

Sixteen DNA microsatellites or simple sequence length polymorphisms (SSLPs), generated by polymerase chain reaction (PCR) were selected for use in the genetic quality control of the nine inbred SENCAR strains currently available. The SENCAR strains constitute a powerful tool for mechanistic studies of multi-stage skin carcinogenesis, as well as for studies to understand the underlying genetic basis of resistance to tumour promotion and progression. SSLP analysis is a fast and economical way for detecting genetic contamination (unexpected outcrosses) among these closely-related albino strains, where standard immunological and biochemical markers have been shown to be insufficient.     

Representation of amino acids as five-bit or three-bit patterns for filtering protein databases

MOTIVATION: We propose representing amino acids by bit-patterns so they may be used in a filter algorithm for similarity searches over protein databases, to rapidly eliminate non-homologous regions of database sequences. The filter algorithm would be based on dynamic programming optimization. It would have the advantage over previous filter algorithms that its substitution scoring function distinguishes between conservative and non-conservative amino acid substitutions. RESULTS: Simulated annealing was used to search for the best five-bit or three-bit patterns to represent amino acids, where similar amino acids were given similar bit-patterns. The similarity between amino acids was estimated from the BLOSUM45 matrix. Representing amino acids by these five-bit and three-bit patterns, the Escherichia coli PhoE precursor and the bacteriophage PA2 LC precursor were aligned. The alignments were nearly the same as that obtained when BLOSUM45 was used to score substitutions. AVAILABILITY: The C code of the optimization algorithm for searching for the optimal bit-pattern representation of amino acids is available from the authors upon request.     

Efficient synthesis of anacardic acid analogues and their antibacterial activities

Anacardic acid derivatives exhibit a broad range of biological activities. In this report, an efficient method for the synthesis of anacardic acid derivatives was explored, and a small set of salicylic acid variants synthesised retaining a constant hydrophobic element (a naphthyl tail). The naphthyl side chain was introduced via Wittig reaction and the aldehyde installed using directed ortho-metalation reaction of the substituted o-anisic acids. The failure of ortho-metalation using unprotected carboxylic acid group compelled us to use directed ortho-metalation in which a tertiary amide was used as a strong ortho-directing group. In the initial route, tertiary amide cleavage during final step was challenging, but cleaving the tertiary amide before Wittig reaction was beneficial. The Wittig reaction with protected carboxylic group (methyl ester) resulted in side-products whereas using sodium salt resulted in higher yields. The novel compounds were screened for antibacterial activity and cytotoxicity. Although substitution on the salicylic head group enhanced antibacterial activities they also enhanced cytotoxicity.