Patterns of junctional communication in skin: Studies on cultured keratinocytes

Junctional communication has long been suggested to play a role in coordinating the development of multicellular tissues. A better understanding of the patterns of communication between cells in such tissues is important for the identification of areas where this process may have a role. We have investigated the patterns of communication in cultures of human epidermal keratinocytes by iontophoretic injection of Lucifer Yellow CH, using involucrin expression as a marker of cells undergoing terminal differentiation. Cells that lack involucrin (i.e., the basal, proliferating cells) transfer dye preferentially to other involucrin-negative cells, whereas involucrin-positive cells either are not coupled or transfer dye with similar frequency to involucrin-positive and involucrin-negative neighbors. This decrease in communication associated with terminal differentiation was observed in both the presence and the absence of assembled desmosomes. Our observations lead us to speculate that loss of junctional communication may influence the commitment of basal keratinocytes to terminal differentiation.     

Siliceous protozoan plates in lake sediments

The silicified plates of certain testate amoebae have the necessary characteristics to be useful paleoindicators. These plates are usually abundant and well preserved in sedimentary deposits, where they can be studied using the same preparation techniques developed for other siliceous microfossils. Individual plates can usually be identified to the generic and, in some cases, to the species level. A preliminary survey of plates recorded in the surficial (top centimetre) sediments of 38 Adirondack Mountain (NY) lakes suggests that these testaceans may reflect catchment features (e.g. bog development) more closely than lakewater characteristics. The siliceous, species-specific scales of heliozoans are also shown to preserve in sedimentary profiles.     

The biogeography of AustralianDaphnia: clues of an ancient (>70 m.y.) origin for the genus

The AustralianDaphnia (D. occidentalis, D. jollyi, D. lumholtzi, D. carinata, D. nivalis, andD. cephalata) include representatives from three distinct groups within the genus. The worldwide distribution of species within these three groups is consistent with a Gondwanan or pre-Gondwanan origin for each group. These data suggest an even more ancient origin for the genusDaphnia. The finding thatD. cephalata must have been separated fromD. carinata since Gondwanan times is consistent with recent biochemical data on the phylogenetic relations of AustralianDaphnia, but contrasts with electrophoretic evidence of continuing hybridization between these two taxa. It is suggested that continued gene exchange between otherwise discrete species, or clonal complexes, ofDaphnia may be maintained for millions of years, partly as a result of the varying degrees of cyclical parthenogenesis, and partly as a result of the highly fragmented population structures, found in these taxa.     

An analysis of the organization and evolution of type 4 fimbrial (MePhe) subunit proteins

Summary We have analyzed and compared the amino acid sequences of the type 4 fimbrial subunits fromPseudomonas aeruginosa, Moraxella bovis, M. nonliquefaciens, Bacteroides nodosus, Neisseria gonorrhoeae, andN. meningitidis. We propose a consensus sequence for the highly conserved aminoterminal regions of these proteins. In the variable regions, a domain corresponding to an epitope common toN. gonorrhoeae andN. meningitidis fimbriae is conserved, both in sequence and in environment, in fimbrial subunits fromB. nodosus. The subunits fromM. bovis andP. aeruginosa do not show any homologies to this sequence. In all of the subunits, the carboxy-terminal half of the molecule consists of a series of fairly hydrophobic domains. The last three domains, two of which include the cysteines of the disulfide bridge inN. gonorrhoeae, P. aeruginosa, andM. bovis, are more or less conserved in sequence in all of the proteins including that ofB. nodosus. We propose that these conserved hydrophobic regions, which have the potential to form a series of beta-sheets, form a structural framework around which more variable hydrophilic sequences determining immunological profile are arranged. The evolutionary relationships of the contemporary proteins and the distribution of type 4 fimbriae are also discussed.     

The genetic relationships between the kringle domains of human plasminogen,prothrombin, tissue plasminogen activator,urokinase, and coagulation factor XII

Summary A computer-based statistical evaluation of the optimal alignments of the kringle domains of human plasminogen, human prothrombin, human tissue plasminogen activator, human urokinase, and human coagulation Factor XIIa, as well as the putative kringle of human haptoglobin, has been performed. A variety of different alignments has been examined and scores calculated in terms of the number of standard deviations (SD) of a given match from randomness. With the exception of human haptoglobin, it was found that very high alignment scores (8.9–23.0 SD from randomness) were obtained between each of the kringles, with the kringle 1 and kringle 5 regions of human plasminogen displaying the highest similarity, and the S kringle of human prothrombin and the human Factor XII kringle showing the least similarity. The relationships obtained were employed to construct an evolutionary tree for the kringles. The predicted alignments have also allowed nucleotide mutations in these regions to be evaluated more accurately. For those regions for which nucleotide sequences are known, we have employed the maximal alignments from the protein sequences to assess nucleotide sequence similarities. It was found that a range of approximately 40–55% of the nucleotide bases were placed at identical positions in the kringles, with the highest number found in the alignment of the two kringles of human tissue plasminogen activator and the lowest number in the alignment of the S kringle of prothrombin with the second kringle of tissue plasminogen activator. From both protein and nucleotide alignments, we conclude that haptoglobin is not statistically homologous to any other kringle.Secondary structural comparisons of the kringle regions have been predicted by a combination of the Burgess and Chou-Fasman methods. In general, the kringles display a very high number of -turns, and very low -helical contents. From analysis of the predicted structures in relationship to the functional properties of these domains, it appears as though many of their functional differences can be related to possible conformational alterations resulting from amino acid substitutions in the kringles.     

Time Course of Adaptations in Dopamine Biosynthesis, Metabolism, and Release Following Nigrostriatal Lesions: Implications for Behavioral Recovery from Brain Injury

Alterations in neostriatal dopamine metabolism, release, and biosynthesis were determined 3, 5, or 18 days following partial, unilateral destruction of the rat nigrostriatal dopamine projection. Concentrations of dopamine and each of its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 3-methoxytyramine (3-MT) were markedly decreased in the lesioned striata at 3, 5, or 18 days postoperation. The decline in striatal high-affinity [3H]dopamine uptake closely matched the depletion of dopamine at 3 and 18 days postoperation. However, neither DOPAC, HVA, nor 3-MT concentrations were decreased to as great an extent as dopamine at any time following lesions that depleted the dopamine innervation of the striatum by greater than 80%. In these more severely lesioned animals, dopamine metabolism, estimated from the ratio of DOPAC or HVA to dopamine, was increased two- to four-fold in the injured hemisphere compared with the intact hemisphere. Dopamine release, estimated by the ratio of 3-MT to dopamine, was more increased, by five- to sixfold. Importantly, the HVA/dopamine, DOPAC/dopamine, and 3-MT/dopamine ratios did not differ between 3 and 18 days postlesioning. The rate of in vivo dopamine biosynthesis, as estimated by striatal DOPA accumulation following 3,4-dihydroxyphenylalanine (DOPA) decarboxylase inhibition with NSD 1015, was increased by 2.6- to 2.7-fold in the surviving dopamine terminals but again equally at 3 and 18 days postoperation. Thus, maximal increases in dopamine metabolism, release, and biosynthesis occur rapidly within neostriatal terminals that survive a lesion. This mobilization of dopaminergic function could contribute to the recovery from the behavioral deficits of partial denervation by increasing the availability of dopamine to neostriatal dopamine receptors. However, these presynaptic compensations are not sufficient to account for the protracted (at least 3-week) time course of sensorimotor recovery that has been observed following partial nigrostriatal lesion.     

N1-Methyl-2-125I-Lysergic Acid Diethylamide, a Preferred Ligand for In Vitro and In Vivo Characterization of Serotonin Receptors

Methylation of 2-125I-lysergic acid diethylamide (125I-LSD) at the N1 position produces a new derivative, N1-methyl-2-125I-lysergic acid diethylamide (125I-MIL), with improved selectivity and higher affinity for serotonin 5-HT2 receptors. In rat frontal cortex homogenates, specific binding of 125I-MIL represents 80-90% of total binding, and the apparent dissociation constant (KD) for serotonin 5-HT2 receptors is 0.14 nM (using 2 mg of tissue/ml). 125I-MIL also displays a high affinity for serotonin 5-HT1C receptors, with an apparent dissociation constant of 0.41 nM at this site. 125I-MIL exhibits at least 60-fold higher affinity for serotonin 5-HT2 receptors than for other classes of neurotransmitter receptors, with the dopamine D2 receptor as its most potent secondary binding site. Studies of the association and dissociation kinetics of 125I-MIL reveal a strong temperature dependence, with very slow association and dissociation rates at 0 degree C. Autoradiographic experiments confirm the improved specificity of 125I-MIL. Selective labeling of serotonin receptors was observed in all brain areas examined. In vivo binding studies in mice indicate that 125I-MIL is the best serotonin receptor label yet described, with the highest frontal cortex to cerebellum ratio of any serotonergic radioligand. 125I-MIL is a promising ligand for both in vitro and in vivo labeling of serotonin receptors in the mammalian brain.     

Genetic transformation of flax (Linum usitatissimum) by Agrobacterium tumefaciens: regeneration of transformed shoots via a callus phase

Genetic transformation of flax (Linum usitatissimum) has been achieved using an A. tumefaciens strain carrying a non-oncogenic Ti plasmid-derived vector containing a chimaeric npt-II gene and a wild type nopaline synthase gene. Fertile, transformed shoots were most easily obtained from Km^r callus developing on hypocotyl sections. The totipotency of the Km^r callus was dependent upon its origin. T-DNA was visualised by Southern blotting in all Km^r tissues. Efficient expression of nopaline synthase and the chimaeric npt-II gene was found in transformed Km^r callus and regenerated shoots.Abbreviations npt-II neomycin phosphotransferase II gene - NPT-II neomycin phosphotransferase II - nos nopaline synthase gene promoter - Km^r kanamycin resistant - BAP 6-benzylaminopurine - NAA -naphthaleneacetic acid - MSD4×2 medium D4×2 based on Murashige & Skoog medium (see Scott & Draper, 1987)     

Sterol conjugates of two phenotypically different calli of Beta vulgaris

Steryl glycosides are the predominant form of sterol at 88% of the total sterol in non-betalain producing calli of Beta vulgaris. The total sterol decreases and sterol form shifts from steryl glycosides to 97% free sterol upon the transition of non-betalain to betalain producing calli. A substantial decrease in stigmasterol (24--ethylcholesta-5,22E-dien-3-ol) and sitosterol (24-ethylcholest-5-en-3-ol) levels is observed during this transition, and alters the ratio of ⁷:⁵ sterols. Spinasterol (24- ethyl-5-cholesta-7,22E-dien-3-ol) is the dominant sterol at 43% and 95% of the total sterol in non-betalain producing and betalain producing calli. The level of 22-dihydrospinasterol (24-ethyl-5-cholest-7-en-3-ol) is reduced in both calli to 3% from 25% in leaves. Lanosterol (4,4,14-trimethyl-cholesta-8(9),24-dien-3-ol) and cycloartenol (9,19-cyclopropyl-4,4,14-trimethyl-cholest-24-en-3-ol) were identified in betalain and nonbetalain producing callus respectively.