Trithorax: A new homoeotic mutation of Drosophila melanogaster causing transformations of abdominal and thoracic imaginal segments

Summary A new homoeotic mutation, trithorax ¹ (trx ¹), of Drosophila melanogaster is described which causes intersegmental transformations in both the adult thorax and abdomen. Specifically, the metathorax and ventral prothorax are partially transformed to mesothroax, whilst abdominal segments are partially transformed towards the first abdominal segment. Three variables are shown to influence the expressivity of the mutant phenotype: (1) presence of wild type copies of the gene in the female parent; (2) culture temperature during the first four hours of embryogenesis; (3) exposure of embryos to ether vapour at the blastoderm stage. In addition it is shown that phenocopies of the mutant phenotype can be induced by ether treatment of heterozygous trx ¹ embryos with a frequency higher than that of bithorax phenocopies in wild type embryos.These findings are together taken to imply that the mutated gene functions early during embryogenests, and that a common mechanism underlies the determination of all the thoracic and abdominal segments.     

Hyperparathyroidism after renal transplantation.

Renal transplantation reduced circulating parathyroid hormone in recipients of renal allografts. Nevertheless, biochemical and histological evidence of mild hyperparathyroidism persisted up to eight years after renal transplantation despite good graft function.     

Simplified Automated Image Analysis for Detection and Phenotyping of Mycobacterium tuberculosis on Porous Supports by Monitoring Growing Microcolonies

Background

Even with the advent of nucleic acid (NA) amplification technologies the culture of mycobacteria for diagnostic and other applications remains of critical importance. Notably microscopic observed drug susceptibility testing (MODS), as opposed to traditional culture on solid media or automated liquid culture, has shown potential to both speed up and increase the provision of mycobacterial culture in high burden settings.

Methods

Here we explore the growth of Mycobacterial tuberculosis microcolonies, imaged by automated digital microscopy, cultured on a porous aluminium oxide (PAO) supports. Repeated imaging during colony growth greatly simplifies “computer vision” and presumptive identification of microcolonies was achieved here using existing publically available algorithms. Our system thus allows the growth of individual microcolonies to be monitored and critically, also to change the media during the growth phase without disrupting the microcolonies. Transfer of identified microcolonies onto selective media allowed us, within 1-2 bacterial generations, to rapidly detect the drug susceptibility of individual microcolonies, eliminating the need for time consuming subculturing or the inoculation of multiple parallel cultures.

Significance

Monitoring the phenotype of individual microcolonies as they grow has immense potential for research, screening, and ultimately M. tuberculosis diagnostic applications. The method described is particularly appealing with respect to speed and automation.     

Effect of EDTA and Methionine on Preventing Loss of Viscosity of Cellulose-Based Topical Gel

Methylcellulose and hydroxypropylmethylcellulose (hypromellose) are used in topical formulations of a protein to form a viscous hydrogel. Five lots of hypromellose raw material were made into 3% gel; all showed viscosity loss after sterilization by autoclave. EDTA (edetate disodium) minimized the viscosity loss caused by autoclaving in the presence of up to 100 ppm H₂O₂. These results suggest that EDTA may prevent loss of viscosity of the hydrogel when peroxide is present. H₂O₂ at low levels (2–50 ppm) caused significant viscosity loss over time at either 40°C or 5°C in 3% methylcellulose or hypromellose gel. EDTA slowed the rate of viscosity loss during storage under stress by H₂O₂ but did not completely prevent the loss. Methionine was effective in completely preventing gel-viscosity loss during storage in the presence of up to 50 ppm H₂O₂. On the basis of these results, it is recommended that methionine be added to the protein topical formulation as a stabilizer against viscosity loss.     

Identification of a novel inhibitory actin-capping protein binding motif in CD2-associated protein

CD2-associated protein (CD2AP) is a scaffold molecule that plays a critical role in the maintenance of the kidney filtration barrier. Little, however, is understood about its mechanism of function. We used mass spectrometry to identify CD2AP-interacting proteins. Many of the proteins that we identified suggest a role for CD2AP in endocytosis and actin regulation. To address the role of CD2AP in regulation of the actin cytoskeleton, we focused on characterizing the interaction of CD2AP with actin-capping protein CP. We identified a novel binding motif LXHXTXXRPK(X)6P present in CD2AP that is also found in its homolog Cin85 and other capping protein-associated proteins such as CARMIL and CKIP-1. CD2AP inhibits the function of capping protein in vitro. Therefore, our results support a role of CD2AP in the regulation of the actin cytoskeleton.     

Growth and Multiplexed Analysis of Microorganisms on a Subdivided, Highly Porous, Inorganic Chip Manufactured from Anopore

A highly porous inorganic material (Anopore) was shown to be an effective support for culturing and imaging a wide range of microorganisms. An inert barrier grid was printed on the rigid surface of Anopore to create a “living chip” of 336 miniaturized compartments (200/cm²) with broad applications in microbial culture.     

Reversible unfolding of the gelatin-binding domain of fibronectin: structural stability in relation to function

Fibronectin, a large multidomain glycoprotein, binds denatured collagen (gelatin) and mediates cell attachment and spreading on collagen-coated surfaces. Despite the high affinity, binding to gelatin is disrupted by relatively mild conditions. We have examined the effects of denaturants on the structure and function of a 42-kDa gelatin-binding fragment (GBF) isolated from chymotryptic and thermolytic digests of the parent protein. Application of linear gradients to GBF-loaded gelatin-agarose columns resulted in peak elution of the fragment at pH 5.2 or 10.2, at 0.4 M dimethylformamide, 0.9 M GdmCl, or 2.0 M urea, conditions far short of those required to induce structural changes detectable by fluorescence or circular dichroism. Solvent perturbation, fluorescence quenching, and chemical modification experiments indicate that about half of the 8 tryptophans, one-third of the 21 tyrosines, and all of the 9 lysine residues are solvent-exposed in the native protein and that 1 or more of the latter are directly involved in binding to gelatin, most likely through a hydrogen-bonding mechanism. Titration with GdmCl produced a single unfolding transition centered near 2.5 M GdmCl as monitored by changes in fluorescence and circular dichroism. This transition was fully reversible with complete recovery of structural parameters and gelatin binding. Treatment with disulfide reducing agents caused rapid irreversible changes in structure similar to those produced by GdmCl with concomitant loss of gelatin binding. Thus, tertiary and secondary structures are important for binding, but binding can be disrupted without perturbing those structures.     

Bacteriological study of otogenic cerebral abscesses: chemotherapeutic role of metronidazole.

In nine consecutive patients with otogenic cerebral abscesses a mixed growth of aerobic and obligate anaerobic bacteria was isolated from the pus in five patients, and in the remaining four obligate anaerobes were the sole isolates. The commonest obligate anaerobe isolated was Bacteroides fragilis, which was present in all but one patient. The patients were all treated with metronidazole for the anaerobic organisms and with appropriate chemotherapy against the aerobic organisms isolated. All the patients recovered and only one was left with a neurological deficit. As otogenic cerebral abscesses constitute a major proportion of all cerebral abscesses, the use of metronidazole against obligate anaerobic bacteria, which tend to dominate in such abscesses, should reduce the high mortality from this condition.