Localization of sequences regulating ancestral and acquired sites of esterase6 activity in Drosophila melanogaster

We have broadly defined the DNA regions regulating esterase6 activity in several life stages and tissue types of D. melanogaster using P- element-mediated transformation of constructs that contain the esterase6 coding region and deletions or substitutions in 5' or 3' flanking DNA. Hemolymph is a conserved ancestral site of EST6 activity in Drosophila and the primary sequences regulating its activity lie between -171 and -25 bp relative to the translation initiation site: deletion of these sequences decrease activity approximately 20-fold. Hemolymph activity is also modulated by four other DNA regions, three of which lie 5' and one of which lies 3' of the coding region. Of these, two have positive and two have negative effects, each of approximately twofold. Esterase6 activity is present also in two male reproductive tract tissues; the ejaculatory bulb, which is another ancestral activity site, and the ejaculatory duct, which is a recently acquired site within the melanogaster species subgroup. Activities in these tissues are at least in part independently regulated: activity in the ejaculatory bulb is conferred by sequences between -273 and -172 bp (threefold decrease when deleted), while activity in the ejaculatory duct is conferred by more distal sequences between -844 and -614 bp (fourfold decrease when deleted). The reproductive tract activity is further modulated by two additional DNA regions, one in 5' DNA (-613 to -284 bp; threefold decrease when deleted) and the other in 3' DNA (+1860 to +2731 bp; threefold decrease when deleted) that probably overlaps the adjacent esteraseP gene. Collating these data with previous studies suggests that expression of EST6 in the ancestral sites is mainly regulated by conserved proximal sequences while more variable distal sequences regulate expression in the acquired ejaculatory duct site.      

A lipoprotein signal peptide encoded by the staphylococcal conjugative plasmid pSK41 exhibits an activity resembling that of Enterococcus faecalis pheromone cAD1.

The traH gene of the staphylococcal conjugative plasmid pSK41 has recently been shown to encode a lipoprotein (N. Firth, K. P. Ridgway, M. E. Byrne, P. D. Fink, L. Johnson, I. T. Paulsen, and R. A. Skurray, Gene 136:13-25, 1993). Here we report that traH encodes a product recognized as a pheromone by Enterococcus faecalis cells harboring the conjugative plasmid pAD1. The mature traH product is not essential for this phenomenon, as expression of pheromone-like activity was found to require sequences encoding only the pro-TraH signal peptide.     

Phylogenetic diversification of immunoglobulin genes and the antibody repertoire

Immunoglobulins are encoded by a large multigene system that undergoes somatic rearrangement and additional genetic change during the development of immunoglobulin-producing cells. Inducible antibody and antibody-like responses are found in all vertebrates. However, immunoglobulin possessing disulfide-bonded heavy and light chains and domain-type organization has been described only in representatives of the jawed vertebrates. High degrees of nucleotide and predicted amino acid sequence identity are evident when the segmental elements that constitute the immunoglobulin gene loci in phylogenetically divergent vertebrates are compared. However, the organization of gene loci and the manner in which the independent elements recombine (and diversify) vary markedly among different taxa. One striking pattern of gene organization is the "cluster type" that appears to be restricted to the chondrichthyes (cartilaginous fishes) and limits segmental rearrangement to closely linked elements. This type of gene organization is associated with both heavy- and light-chain gene loci. In some cases, the clusters are "joined" or "partially joined" in the germ line, in effect predetermining or partially predetermining, respectively, the encoded specificities (the assumption being that these are expressed) of the individual loci. By relating the sequences of transcribed gene products to their respective germ-line genes, it is evident that, in some cases, joined-type genes are expressed. This raises a question about the existence and/or nature of allelic exclusion in these species. The extensive variation in gene organization found throughout the vertebrate species may relate directly to the role of intersegmental (V<==>D<==>J) distances in the commitment of the individual antibody-producing cell to a particular genetic specificity. Thus, the evolution of this locus, perhaps more so than that of others, may reflect the interrelationships between genetic organization and function.      

Human cyclins B1 and B2 are localized to strikingly different structures: B1 to microtubules, B2 primarily to the Golgi apparatus.

We have raised and characterized antibodies specific for human cyclin B2 and have compared the properties of cyclins B1 and B2 in human tissue culture cells. Cyclin B1 and B2 levels are very low in G1 phase, increase in S and G2 phases and peak at mitosis. Both B-type cyclins associate with p34cdc2; their associated kinase activities appear when cells enter mitosis and disappear as the cyclins are destroyed in anaphase. However, human cyclins B1 and B2 differ dramatically in their subcellular localization. Cyclin B1 co-localizes with microtubules, whereas cyclin B2 is primarily associated with the Golgi region. In contrast to cyclin B1, cyclin B2 does not relocate to the nucleus at prophase, but becomes uniformly distributed throughout the cell. The different subcellular locations of human cyclins B1 and B2 implicate them in the reorganization of different aspects of the cellular architecture at mitosis and indicate that different mitotic cyclin-cyclin-dependent kinase complexes may have distinct roles in the cell cycle.     

A comparison of the effects of different perfusion regimes on the structure of the isolated human placental lobule

Summary Isolated lobules of normal term human placentas were perfused using two different procedures. In the first more conventional system, open-circuit perfusion of both the maternal and the fetal circulations with Earle's solution containing dextran was established and maintained for either 30 min or 1 h. In the second series of experiments both circulations were perfused in separate closed circuits with a mixture of fresh autologous fetal blood and Earle's solution for 0, 1, 2 or 3 h. In both series the lobule was then fixed by perfusion through the fetal circulation.Light and electron-microscopic examination of a set of tissue samples from each perfused lobule showed substantial differences between the effects of these two types of perfusion procedure. Tissue from lobules perfused by the open-circuit blood-free procedure showed patchy but severe cell swelling and vacuolation of the trophoblast after only one hour's perfusion. Particularly striking was swelling and disruption of a large proportion of the mitochondria in all placental cell types. By contrast, placental tissue from the closed-circuit perfusion with blood-containing medium showed little change over a period of two hours, while after three hours it showed oedema and microvillous damage, but no sign of cell swelling and little mitochondrial damage.It is concluded that the viability of the perfused human placental lobule depends on the type of perfusate used, and that the use of a fetal blood-enriched perfusate is of considerable value in maintenance of the preparation as assessed by structural criteria.     

Ultrastructural study of the permeability of the guinea-pig placenta to horseradish peroxidase

Summary Horseradish peroxidase (HRP) was used to study macromolecule permeation into the guinea-pig placenta perfused in situ. When tissue culture medium 199 (TC 199) was used as fetal-side perfusate, the tracer reaction product was found only lining the fetal endothelium. When a longer period of perfusion with HRP in TC 199 was used, a small amount of reaction product was found in the subendothelial space and syncytiotrophoblastic vesicles, but not in maternal lacunae. In similar experiments using a Krebs bicarbonate Ringer (KRBG) as perfusate the tracer was found (i) lining the fetal endothelium, (ii) in the lateral intercellular spaces of the endothelium, (iii) in the subendothelial space, and (iv) in the maternal lacunae.It is therefore evident that the vehicle influenced the permeability of the guinea-pig placenta to horseradish peroxidase. As other studies have shown that perfusion of the fetal side with salt solution increases pore size, the results with TC 199 are regarded as more representative of the situation in the intact animal. It is therefore suggested that the fetal endothelium of the guinea-pig placenta may be largely impermeable to molecules of the size of horseradish peroxidase (4 nm) or larger.     

Structural features and quantitative age-dependent changes in the intervascular barrier of the guinea-pig haemochorial placenta

Summary The haemomonochorial placenta of the guinea-pig undergoes several quantitative changes between the 49th and 64th days of gestation, all of which are in such a direction as to increase the efficiency of transplacental transport. The fetal vessels become larger, the maternal vessels increase in surface area by proliferation of microvilli, and the effective mean distance between the two vessel sets decreases. The magnitude of these changes suggests that the efficiency of transport of hydrophilic solutes across the maternal-fetal interface could double, although changes in the number of permeation sites per unit area may modify this relationship. The presence of open intercellular spaces and fenestrations in the fetal endothelium suggests that this layer may not be a major permeability barrier in the guinea-pig, but may create an unstirred layer of extracellular fluid between endothelium and syncytiotrophoblast.     

Home accidents in older people: role of primary health care team.

OBJECTIVES--To determine the incidence and nature of unreported and reported home accidents in older people and to investigate associated environmental factors. DESIGN--Postal questionnaire requesting information on home accidents in the preceding month. SETTING--Inner London general practice. SUBJECTS--All registered patients aged over 65 years (n = 1662), of whom 120 were inappropriately registered and 1293 responded. MAIN OUTCOME MEASURE--Circumstances and consequences of accidents in the home. RESULTS--108 accidents were recorded in 100 patients, giving a home accident rate of 84/1000 patients, equivalent to an annual rate of 1002/1000. 73 accidents were falls, and 83 were unreported. Of the 25 reported accidents, 19 were reported to general practice and six to accident and emergency departments (5.6% of all events). Rates of home accidents increased with age and were higher in women than men (79/819 upsilon 29/474; chi 2 = 4.5, df = 1, p less than 0.05). CONCLUSIONS--The incidence of home accidents in people aged over 65 years was high but few events were reported to medical services. General practice provided the main contact for patients who reported home accidents, and primary care workers have important opportunities for advising elderly patients on home accident prevention. Improved publicity on home safety targeted at older people and their carers would support the primary health care team in this role.