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Two protochordate genes encode pituitary adenylate cyclase-activating polypeptide and related family members.

To address the origin of the glucagon superfamily, we isolated and sequenced the complementary DNA and partial gene that encode pituitary adenylate cyclase-activating polypeptide (PACAP) from a protochordate (tunicate), a sister group of the amphioxus and vertebrates, but one that evolved before the amphioxus. This is the first report of any superfamily member sequenced from an invertebrate. Transcription of the tunicate pacap1 gene results in a messenger RNA that is 507 bp. The gene contains 3 exons that encode a signal peptide, GRF-like peptide(1-27), and PACAP(1-27). The tunicate GRF-like peptide has 59% identity with human GRF, whereas the deduced amino acids of tunicate PACAP(1-27) have 96% identity with the ovine, human, and salmon PACAP(1-27) forms. Another complementary DNA clone pacap2 was isolated and shown to contain 4 exons that encode a signal peptide, a cryptic peptide, and two peptides that are clearly members of the glucagon superfamily. One of the peptides has 89% sequence identity to the tunicate PACAP encoded in pacap1. A comparison of the two structurally related PACAP clones, each encoding two peptides on separate exons, shows high inter- and intraexon nucleotide sequence identity. Sequence analysis suggests that an exon duplication followed by a gene duplication was responsible for the origin of the two genes. It is argued that the PACAP gene is derived from the protochordate ancestral genes that led to the vertebrate forms of GRF and PACAP.

Amino Acid Sequence↗

Identification and characterization of the murine and human gene encoding the tuberoinfundibular peptide of 39 residues.

By screening public databases, we identified human and mouse genomic DNA clones that encode the tuberoinfundibular peptide of 39 residues (TIP39). The TIP39 precursor is encoded by at least three exons; a noncoding exon U1, exon 1 encoding residues -61 (initiator methionine) to -19 of the leader sequence, and exon 2 encoding residues -18 to -1 and residues +1 to +39. Secreted human TIP39 is identical to the previously isolated bovine TIP39, whereas mouse TIP39 differs by four amino acids. Phylogenetic analyses suggested that TIP39, PTH, and PTHrP may have evolved from a common ancestor. Synthetic human and mouse TIP39 showed indistinguishable potencies [EC(50): 0.54 (human) vs. 0.74 nM (mouse)] at the human PTH2-receptor stably expressed in LLCPK(1) cells; furthermore, TIP-(9-39) was an inhibitor of cAMP accumulation stimulated by either [Tyr(34)]PTH(1-34)amide or human/bovine TIP39. In the mouse, an approximately 4.5-kb mRNA encoding TIP39 was identified by Northern blot analysis in testis and, less abundantly, in liver and kidney, whereas other tissues revealed additional smaller transcripts. In situ hybridizations revealed TIP39 expression in seminiferous tubuli and several brain regions, including nucleus ruber, nucleus centralis pontis, and nucleus subparafascicularis thalami. Because PTH2 receptor expression was previously shown to be highest in brain, pancreas, and testis, our findings are consistent with the notion that TIP39 is a neuropeptide which may also have a role in spermatogenesis.

Amino Acid Sequence↗

Gene-encoding parathyroid hormone-like peptide: nucleotide sequence of the rat gene and comparison with the human homologue.

The single-copy gene coding for rat PTH-like peptide was isolated from a rat liver genomic DNA library. The gene spans 12 kilobases and contains four exons. Exon I encodes the 5'-noncoding region, exon II encodes the prepro region, exon III encodes the mature peptide sequence up to amino acid 139 and exon IV encodes the carboxyl-terminal two amino acids, a stop codon, and the 3'-noncoding region. Splicing of these exons yields the 1.4 kilobase mRNA which is the predominant transcript observed in the hypercalcemic rat Leydig cell tumor and several normal rat tissues. The overall exon/intron organization of the rat parathyroid hormone-like peptide (PLP) gene is similar to that of the PTH gene and emphasizes the likelihood that PLP and PTH arose from a common ancestral gene. A comparison of the single promoter region of the rat with the second promoter of the human gene indicates conserved TATA and CAAT box homologies, GC box regions (SP-1 binding sites), putative AP-2 binding sites, and a vitamin D responsive element. When compared to the seven exon human PLP gene, which uses multiple promoters and encodes three peptide isoforms, the simpler organization of the rat gene predicts, in mammals, the predominant use of a single promoter and generation of a 141-amino acid peptide as the major molecular form.

Amino Acid Sequence↗

Encoding spatial information in the waggle dance.

Apis mellifera bees execute waggle dances to recruit other bees to desirable food sources. Several components of the waggle dance are correlated with the direction of and the distance to food. Moreover, recruits use the spatial information encoded in the dance to locate the signalled food. However, although dance communication has been studied extensively, little is known about how the dancers combine the compass (direction) and the odometric (distance) information they acquire during the foraging flight. In the present study, we analysed the encoding of spatial information in the waggle dance by manipulating the navigational information provided to dancing bees. To this end, we took advantage of the bees' visually driven odometer. We found that the waggle dance basically encodes information on the distance gauged during the outbound (hive-to-food) flight. However, it does not necessarily refer to a global vector based on path integration of the outbound flight. Whenever the direction connecting the subjective food location and the hive does not match the direction of the global vector, dancers refer to a direction close to that of the shortcut connecting the actual food location and the hive. Moreover, in our experiments, this direction was close to that of the inbound (food-to-hive) flight, indicating that landmark-based information is computed during the inbound flight and that it may strongly affect the encoding of directional information in the waggle dance. Moreover, we found that the bees' experience of the terrain modulates the encoding of spatial information in the waggle dance, suggesting that interactions between path integration and visual landmarks are computed in the context of dance communication.

Animal Communication↗

Cloning of a gene encoding a highly stable endo-beta-1,4-glucanase from Aspergillus niger and its expression in yeast.

A gene encoding an endo-beta-1,4-glucanase, which is highly resistant to high temperature, protease and surfactant treatment, was isolated from Aspergillus niger IFO31125 and designated as eng1. The deduced amino acid sequence encoded by eng1 showed high homology with the sequence of a not-well-characterized cellulase encoded by eglB which has not yet been shown to be a stable enzyme. To confirm the sequence of the gene encoding the highly stable endo-beta-1,4-glucanase, the cloned gene was expressed in the yeast Saccharomyces cerevisiae, in which no cellulase activity was found, and the gene product was purified and subjected to enzymatic characterization. The enzyme retained 56% of the initial activity after 1 h of incubation at 80 degrees C and was stable in the range of pH 3.0-10.0. The optimal temperature for enzyme activity was 70 degrees C and the optimal pH was 6.0. The enzyme was highly protease-resistant and retained more than 80% of the initial activity after protease treatment for 3 d at 40 degrees C. The enzyme was also resistant to various surfactants. From these results, eng1 was confirmed to encode a very stable endo-beta-1,4-glucanase.

Journal Article↗

Modified minimum-distance criterion for blended random and nonrandom encoding.

Two pixel-oriented methods for designing Fourier transform holograms--pseudorandom encoding and minimum-distance encoding-usually produce higher-fidelity reconstructions when combined than those produced by each method individually. In previous studies minimum-distance encoding was defined as the mapping from the desired complex value to the closest value produced by the modulator. This method is compared with a new minimum-distance criterion in which the desired complex value is mapped to the closest value that can be realized by pseudorandom encoding. Simulations and experimental measurements using quantized phase and amplitude modulators show that the modified approach to blended encoding produces more faithful reconstructions than those of the previous method.

Algorithms↗

Detection of chromosome 21-encoded mRNA of placental origin in maternal plasma.

BACKGROUND: mRNA of placental origin (i.e., human placental lactogen and beta-human chorionic gonadotropin) has been demonstrated to be easily detectable in maternal plasma. We tested whether detection of chromosome 21-encoded mRNA of placental origin is possible in maternal plasma obtained during the first trimester. METHODS: Plasma samples were obtained from pregnant women between weeks 9-13 of pregnancy. RNA was isolated from 800 or 1600 microL of plasma by silica-based affinity isolation and, after on-column DNase treatment, was subjected to two-step, one-tube reverse transcription-PCR with gene specific primers. RESULTS: Three chromosome 21-encoded genes located within the Down syndrome critical region with overexpression in trisomy 21 placentas were screened for expression in early placental tissue to select their potential use for RNA based plasma screening. One of the chromosome 21-encoded genes (LOC90625) showed strong expression in first trimester placenta similar to CSH1 (human placental lactogen) and was selected for plasma analysis. The RNA isolation assay was validated with CSH1 mRNA, which could be detected in the plasma of all women tested in weeks 9-13 of pregnancy. RNA from the chromosome 21-encoded, placentally expressed gene, LOC90625, was present in maternal first-trimester plasma and could be detected in 60% of maternal plasma samples when 800 microL of plasma was used and in 100% of samples when 1600 microL of plasma was used. CONCLUSION: The detection of chromosome 21-encoded mRNA of placental origin in maternal plasma during the first trimester may allow development of plasma-RNA-based strategies for prenatal prediction of Down syndrome. LOC90625 is a candidate gene for this purpose.

Chromosomes, Human, Pair 21↗

Concentration of mRNA encoding 3 beta-hydroxysteroid dehydrogenase/delta 5,delta 4 isomerase (3 beta-HSD) and 3 beta-HSD enzyme activity following treatment of ewes with prostaglandin F2 alpha.

The objectives of these experiments were (1) to determine if prostaglandin F2 alpha (PGF2 alpha) decreased mRNA encoding 3 beta-hydroxysteroid dehydrogenase/d5,delta 4 isomerase (3 beta-HSD) specifically in large steroidogenic luteal cells, which contain the high affinity receptors for PGF2 alpha; and (2) to determine if the decreased concentration of mRNA encoding 3 beta-HSD following administration of PGF2 alpha was associated with a decrease in 3 beta-HSD enzyme activity. Ewes on days 11 or 12 of the estrous cycle were administered PGF2 alpha (25 mg i.v. followed by 10 mg i.m. 2 h later) and corpora lutea collected 4, 12, 24, or 48 h later (n = 4-5/time). Corpora lutea were also collected from non-injected (n = 4) or saline-injected (n = 4) control ewes. Administration of PGF2 decreased (P < 0.05) steady-state concentrations of mRNA encoding 3 beta-HSD to 35, 15, 9, and 5 percent of the concentrations in the control group at 4, 12, 24, and 48 h, respectively. Concentrations of mRNA encoding 3 beta-HSD in large luteal cells were decreased to 43% of controls 4 h following injection, which was similar to the decrease seen in steady-state concentrations of this mRNA in total luteal mRNA (35%). However, 3 beta-HSD enzyme activity was not significantly decreased by 48 h after PGF2 alpha injection. Thus, the dramatic decreased in mRNA encoding 3 beta-HSD was not associated with an immediate decrease in 3 beta-HSD enzyme activity and, therefore, does not appear to be responsible for the acute decrease in secretion of progesterone from ovine luteal tissue during PGF2 alpha-induced luteolysis.

Animals↗

Encoding-specific effects of social cognition on the neural correlates of subsequent memory.

To examine whether social cognition recruits distinct mental operations, we measured brain activity during social ("form an impression of this person") and relatively nonsocial ("remember the order in which person information is presented") orienting tasks. Extending previous research on the neural basis of social cognition, the impression formation task differentially engaged an extensive region of the dorsomedial prefrontal cortex (PFC). In contrast, the nonsocial sequencing task differentially engaged the superior frontal and parietal gyri, precentral gyrus, and the caudate. In addition, we compared encoding activations for subsequently remembered (i.e., hits) to subsequently forgotten (i.e., misses) items. The brain regions in which the blood oxygenation level-dependent signal distinguished subsequent hits from subsequent misses depended on which orienting task was performed at encoding: subsequent memory was correlated with encoding activity only in the medial PFC for impression formation trials but in the right hippocampus for sequencing trials. These data inform two interrelated cognitive issues. First, results underscore the neuroanatomical distinctiveness of social cognition and suggest that previous psychological theories may have neglected important functional differences in how the human brain instantiates social and nonsocial cognitive processes. Second, by demonstrating that activity in different brain regions correlates with subsequent memory as a function of the orienting task performed at encoding, these data provide evidence of the neural basis for encoding specificity, the principle that memory is critically determined by the cognitive process engaged by the initial study episode.

Adult↗

Distinct contributions of hippocampal NMDA and AMPA receptors to encoding and retrieval of one-trial place memory.

Allocentric place memory may serve to specify the context of events stored in human episodic memory. Recently, our laboratory demonstrated that, analogous to event-place associations in episodic memory, rats could associate, within one trial, a specific food flavor with an allocentrically defined place in an open arena. Encoding, but not retrieval, of such flavor-place associations required hippocampal NMDA receptors; retrieval depended on hippocampal AMPA receptors. This might have partly reflected the contributions of these receptors to encoding and retrieval of one-trial place, rather than flavor-place, memory. Therefore, the present study developed a food-reinforced arena paradigm to study encoding and retrieval of one-trial allocentric place memory in rats; memory relied on visuospatial information and declined with increasing retention delay, still being significant after 6 h, the longest delay tested (experiments 1 and 2). Hippocampal infusion of the NMDA receptor antagonist d-AP-5 blocked encoding without affecting retrieval; hippocampal infusion of the AMPA receptor antagonist CNQX impaired retrieval (experiment 3). Finally, we confirmed that the d-AP-5 infusions selectively blocked induction of long-term potentiation, a form of synaptic plasticity, whereas CNQX impaired fast excitatory transmission, at perforant-path dentate gyrus synapses in the dorsal hippocampus in vivo (experiment 4). Our results support that encoding, but not retrieval, of one-trial allocentric place memory requires the NMDA receptor-dependent induction of hippocampal synaptic plasticity, whereas retrieval depends on AMPA receptor-mediated fast excitatory hippocampal transmission. The contributions of hippocampal NMDA and AMPA receptors to one-trial allocentric place memory may be central to episodic memory and related episodic-like forms of memory in rats.

2-Amino-5-phosphonovalerate↗

Neuroanatomical dissociation of encoding processes related to priming and explicit memory.

Priming is a facilitation of cognitive processing with stimulus repetition that can occur without explicit memory. Whereas the functional neuroanatomy of perceptual priming at retrieval is established, encoding processes that initiate priming and explicit memory have not yet been anatomically separated, and we investigated them using event-related functional magnetic resonance imaging. Activations predicting later explicit memory occurred in the bilateral medial temporal lobe (MTL) and left prefrontal cortex (PFC). Activity predicting later priming did not occur in these areas, but rather in the bilateral extrastriate cortex, left fusiform gyrus, and bilateral inferior PFC, areas linked with stimulus identification. Surprisingly, these regions showed response reductions. Our results demonstrate that priming and explicit memory have distinct functional neuroanatomies at encoding, with MTL activations being specific for explicit memory, and suggest that priming is initiated by sharpness of neural responding in stimulus identification areas, consistent with recent electrophysiological evidence regarding priming-related neural oscillations at encoding. We tentatively suggest that this sharpened responding at encoding may set the stage for increased neural processing efficiency at retrieval, with these different neural mechanisms both leading to observed priming-related hemodynamic decreases, and argue that neural measurements at encoding, and not just at retrieval, will be critical in resolving the debate about the neural mechanisms of learning that underlie priming.

Adolescent↗

Neural correlates of relational memory: successful encoding and retrieval of semantic and perceptual associations.

Using event-related functional magnetic resonance imaging, we identified brain regions involved in successful relational memory (RM) during encoding and retrieval for semantic and perceptual associations or in general, independent of phase and content. Participants were scanned while encoding and later retrieving associations between pairs of words (semantic RM) or associations between words and fonts (perceptual RM). Encoding success activity (ESA) was identified by comparing study-phase activity for items subsequently remembered (hits) versus forgotten (misses) and retrieval success activity (RSA) by comparing test-phase activity for hits versus misses. The study yielded three main sets of findings. First, ESA-RSA differences were found within the medial temporal lobes (MTLs) and within the prefrontal cortex (PFC). Within the left MTL, ESA was greater in the anterior hippocampus, and RSA was greater in the posterior parahippocampal cortex/hippocampus. This finding is consistent with the notion of an encoding-retrieval gradient along the longitudinal MTL axis. Within the left PFC, ESA was greater in ventrolateral PFC, and RSA was greater in dorsolateral and anterior PFC. This is the first evidence of a dissociation in successful encoding and retrieval activity within left PFC. Second, consistent with the transfer-appropriate processing principle, some ESA regions were reactivated during RSA in a content-specific manner. For semantic RM, these regions included the left ventrolateral PFC, whereas for perceptual RM, they included occipitoparietal and right parahippocampal regions. Finally, only one region in the entire brain was associated with RM in general (i.e., for both semantic and perceptual ESA and RSA): the left hippocampus. This finding highlights the fundamental role of the hippocampus in RM.

Adolescent↗

Persistence of parahippocampal representation in the absence of stimulus input enhances long-term encoding: a functional magnetic resonance imaging study of subsequent memory after a delayed match-to-sample task.

Recent theoretical models based on cellular processes in parahippocampal structures show that persistent neuronal spiking in the absence of stimulus input is important for encoding. The goal of this study was to examine in humans how sustained activity in the parahippocampal gyrus may underlie long-term encoding as well as active maintenance of novel information. The relationship between long-term encoding and active maintenance of novel information during brief memory delays was studied using functional magnetic resonance imaging (fMRI) in humans performing a delayed matching-to-sample (DMS) task and a post-scan subsequent recognition memory task of items encountered during DMS task performance. Multiple regression analyses revealed fMRI activity in parahippocampal structures associated with the active maintenance of trial-unique visual information during a brief memory delay. In addition to a role in active maintenance, we found that the subsequent memory for the sample stimuli as measured by the post-scan subsequent recognition memory task correlated with activity in the parahippocampal gyrus during the delay period. The results provide direct evidence that encoding mechanisms are engaged during brief memory delays when novel information is actively maintained. The relationship between active maintenance during the delay period and long-term subsequent memory is consistent with current theoretical models and experimental data that suggest that long-term encoding is enhanced by sustained parahippocampal activity.

Adolescent↗

Localization of mRNA encoding c-kit during the initiation of folliculogenesis in ovine fetal ovaries.

The c-kit protein is a transmembrane tyrosine kinase receptor that binds the growth factor stem cell factor. Mutant alleles of the genes coding for both the receptor (c-kit) and its ligand (stem cell factor) affect gametogenesis, development of melanoblasts and some aspects of haematopoiesis. The aim of this study was to examine expression of the c-kit gene during folliculogenesis in fetal sheep ovaries using in situ hybridization. A 422 bp cDNA encoding the extracellular domain of the c-kit protein was amplified from sheep ovarian RNA using reverse-transcription-polymerase chain reaction (RT-PCR), cloned and sequenced. Riboprobes transcribed from the ovine cDNA encoding c-kit were used to detect the presence of mRNA encoding c-kit within the ovaries of fetal sheep on days 90, 100, 120 and 135 of gestation (term = 147 days). In day 90 and 100 fetal ovaries, mRNA encoding c-kit was not detected in association with oogonia during the period of meiosis (to prophase I) but was present in some of the isolated oocytes. In ovaries from day 90 to day 135, mRNA encoding c-kit was detected in the oocytes at every stage of follicular growth--primordial through to antral follicles. This pattern of localization is consistent with that demonstrated in mice.

Animals↗

Expression of mRNA encoding decidualization-associated protein, a variant of acute-phase alpha 2 macroglobulin, by rat uterine tissues during pregnancy and pseudopregnancy.

The patterns of expression of mRNA encoding decidualization-associated protein (DAP) in the rat uterus from early, mid-, and late stages of pregnancy were examined by in situ hybridization to provide an insight into the function of DAP during pregnancy. Parallel studies were performed on rat uteri during the oestrous cycle and oil-induced deciduomata. DAP transcripts were absent in virgin uteri and during the day of implantation (day 5), low in early gestation tissues (days 8, 9) and high during midgestation (days 10-14). Expression of mRNA encoding DAP was first detected on day 8 of pregnancy in decidual cells of the lateral decidual glycogen wing areas. On day 9, mRNA encoding DAP was found in the same area as on day 8 as well as in the outer layer of cells in the antimesometrial decidua adjacent to the myometrium termed the fibrinoid capsula. On days 11 and 12, expression appeared in the decidual cells of the mesometrial decidua, and was particularly strong around the ectoplacental cone and blood vessels that were invaded by the trophoblast. By day 13 and throughout the remainder of pregnancy, expression occurred in the mesometrial decidua and metrial gland. No signal was observed in granulated metrial gland cells. mRNA encoding DAP was also found in the smooth circular muscle coat adjacent to the mesometrial decidua. A similar pattern and cellular localization of expression of mRNA encoding DAP expression was detected during the development of the deciduomata, artificially induced in the absence of a conceptus. The spatial and temporal patterns of expression correlated with a possible role for DAP in mediating fetal-maternal interactions and the establishment of the placental structure.

Animals↗

Association of expression of mRNA encoding the PGF2 alpha receptor with luteal cell apoptosis in ovaries of pseudopregnant mice.

The relationship between the expression of mRNA encoding the prostaglandin F2 alpha (PGF2 alpha) receptor and luteal cell apoptosis as determined by oligonucleosome formation was determined in mouse corpora lutea on days 2 (early phase), 6 (mid-phase), and 11 and 13 (late phase) of pseudopregnancy. No signals for mRNA encoding the PGF2 alpha receptor were detected in the ovary as shown by RNA blot analysis of pregnant mares' serum gonadotrophin (PMSG)-treated mice. The expression of the mRNA that could be detected in the corpora lutea on day 2 after hCG treatment was low, but it was increased on day 6 and reached a plateau on days 11 and 13. On days 11 and 13, in situ hybridization signals for mRNA encoding the PGF2 alpha receptor were localized to large luteal cells of the corpora lutea, especially the cells in the superficial layer. However, ethidium bromide staining revealed marked oligonucleosome formation in the corpora lutea on days 11 and 13. Similarly, positive signals of in situ nick-DNA-end labelling were also detected in the corpora lutea on days 11 and 13. Analysis of the adjacent sections of the corpora lutea on day 13 showed that both signals for DNA strand breaks and mRNA encoding the PGF2 alpha receptor were co-expressed in the corpus luteum. These results suggest that apoptosis during structural luteolysis closely associates with the increased expression of mRNA encoding the PGF2 alpha receptor in luteal cells of pseudopregnant mice.

Animals↗

Effect of oestradiol on mRNA encoding GnRH receptor in pituitary tissue of orchidectomized sheep passively immunized against GnRH.

The effect of oestradiol on tissue concentrations of GnRH receptor and mRNA encoding GnRH receptor was assessed in orchidectomized sheep (wethers) made deficient in GnRH by passive immunization. Wethers were assigned to one of four groups (n = 6 animals per group). Animals in groups 2 and 4 received ovine anti-GnRH sera (200 ml; i.v.) at passive immunization, while antisera against the carrier protein was administered to wethers in groups 1 and 3. Oestradiol was delivered as a continuous infusion (2 micrograms h-1) to wethers in groups 1 and 2. Animals in groups 3 and 4 were infused with vehicle alone. Anterior pituitary tissue was collected at the end of the 48 h infusion. Anti-GnRH sera induced a rapid reduction in the serum concentration of LH. Continuous delivery of oestradiol resulted in a twofold increase (P < 0.05) in tissue concentration of GnRH receptor. This oestradiol-induced response was manifest even in wethers in which endogenous GnRH had been neutralized by passive immunization. Conversely, infusion of oestradiol increased (P < 0.05), and intravenous administration of anti-GnRH sera decreased (P < 0.05), concentrations of mRNA encoding GnRH receptor in pituitary tissue. When delivered in combination, anti-GnRH sera reduced (P < 0.05), but did not eliminate, the oestradiol-induced augmentation of steady-state concentrations of mRNA encoding GnRH receptor. These data demonstrate that the basal concentration of mRNA encoding GnRH receptor is dependent on continued GnRH stimulation. In contrast, the oestradiol-induced increase in steady-state concentration of mRNA encoding GnRH receptor is manifest even in the absence of GnRH.

Animals↗

Multiplexed molecular detection using encoded microparticles and nanoparticles.

Signal-encoded microparticles and nanoparticles have been used to label many reactions simultaneously for target identification in assays, and thus are an indispensable part of multiplex technologies. With the increasing demand for multiplexed molecular detection, encoded particles have evolved from pattern encoding to signal-intensity encoding, and also from signal-molecule encapsulation to signal-molecule tagging. The fabrication and utilization of such nano- and microparticles should advance multiplexed analysis. This short review focuses on how these encoded particles work and briefly touches on their applications in multiplexed molecular detection.

Biological Assay↗