Expression of protein tyrosine phosphatase genes during oogenesis in Drosophila melanogaster.

The spatial and temporal expression of seven Drosophila protein tyrosine phosphatase genes during oogenesis was examined by whole mount in-situ hybridization of antisense RNA probes to ovaries. Our observations indicate diverse expression patterns consistent with multiple roles for protein tyrosine phosphatases in the ovary. DPTP99A and corkscrew transcripts are expressed in follicle cells, consistent with possible roles in the EGF receptor signaling pathway. Transcripts from corkscrew and DPTP10D are detected in the germline during oogenesis and localized to the oocyte during egg chamber development. Localization of the two transcripts is disrupted by mutations in egalitarian and Bicaudal D. DLAR and DPTP4E transcripts are found in the germline during the same developmental stages as DPTP10D transcripts, but their transcripts are not localized to the oocyte. DPTP61F transcription is detected only after stage 6 of oogenesis. After stage 10B these transcripts are transported to the oocyte; thus ovarian transcription of DPTP61F may reflect a maternal contribution of the mRNA for later use during embryogenesis. DPTP69D transcripts are sequestered in the nucleus from stage 7 to stage 10, and then released to the cytoplasm. Our observations suggest that the export of DPTP69D mRNA from the nucleus is temporally regulated during oogenesis.

Cloning and characterization of a new theta-class glutathione-S-transferase (GST) gene, gst-3, from Drosophila melanogaster.

We report here on the cloning and characterization of a new theta-class glutathione-S-transferase (GST) gene, gst-3, from Drosophila melanogaster. Its sequence is distinct from previously characterized Drosophila GST genes, and Southern blotting shows no other closely related genes in the genome. In-situ hybridization localizes the gene to chromosome 2 (55D), near gst-2 (53F), and clearly separate from the gst-D cluster at 87B. The gene is intronless and appears to possess conventional 5' TATA, Cap and 3' polyadenylation signals. A single transcript, approximately 1kb in size, appears to be expressed at high levels in all developmental stages examined. When this gene is overexpressed using various upstream GAL4 driver systems, no striking phenotypes are observed; however, we detect bristle morphology defects in some progeny. The gst-3 gene does not appear to be essential, based upon our observation that mutant flies homozygous for an EP element insertion 5' to the TATA box produce little or no detectable gst-3 mRNA; these flies are viable and fertile at 25 and 29 degrees C. Nevertheless, the gst-3 gene appears to be evolutionarily conserved in other Drosophila species, suggesting that it may be functionally important.

Cellular architecture and topographic organization of the inferior colliculus of the squirrel monkey.

The inferior colliculus of the squirrel monkey is made up of a large central nucleus, bordered by the smaller external and pericentral nuclei. The majority of cells in the central nucleus exhibit a pronounced laminar arrangement due to the orientation of their dendrites. In medial sections of the nucleus these laminae lie in a dorsorostral to ventrocaudal direction. More laterally the layers assume a horizontal orientation and at the far lateral edge of the central nucleus come to lie in a ventrorostral to dorsocaudal orientation. A single tonotopic representation of audible frequencies is present in the central nucleus. A regular progression of best frequencies from low to high is encountered as a microelectrode advances from dorsocaudal to ventrorostral in the sagittal plane. Penetrations in more medial regions of the central nucleus encounter neurons whose best frequencies represent a higher range of frequencies than those in the lateral parts. The orientation of the isofrequency laminae determined physiologically appears congruent with the orientation of the dendritic laminae. The relative volume of the central nucleus devoted to each octave from 250 Hz to 32 kHz was determined. Frequencies up to eight kHz command successively larger amounts of collicular tissue. The octave band from 8 to 16 kHz is represented by the greatest amount of collicular tissue. Disproportionate representation of frequency may be the consequence of innervation density along the basilar membrane.

Projections of auditory cortex upon the thalamus and midbrain in the owl monkey.

Two tonotopically organized cortical fields, the primary (AI) and the rostral (R) fields, comprise the core of auditory cortex in the owl monkey. Injections of tritiated proline were made into each of these fields to determine their efferent projections using autoradiographic methods. Both AI and R project to the principal and magnocellular divisions of the medial geniculate body. In addition, R projects to the posterior part of the dorsal division of the medial geniculate. AI sends axons to the dorsomedial region and laminated portion of the central nucleus of the inferior colliculus. Labeling in the central nucleus following AI injections appears as a band of silver grains oriented parallel to isofrequency contours. Axons from R terminate in the dorsomedial region of the central nucleus of the inferior colliculus and in the pericentral and external nuclei of the inferior colliculus. In addition, the rostral field projects to a small area of the medial pulvinar just anterior to the brachium of the superior colliculus.

Auditory cortico-cortical connections in the owl monkey.

Two tonotopically organized cortical fields, the primary (A1) and rostral (R) fields, comprise a core of auditory cortex in the owl monkey. Injections of tritiated proline were made into each of these fields to determine their projections to the auditory fields in the ipsilateral and contralateral hemispheres using autoradiographic methods. Neurons in R project to the rostromedial (RM) and primary fields in both hemispheres, and to the posterolateral (PL) and anterolateral (AL) fields in the ipsilateral hemisphere. In addition, the rostral fields in the two hemispheres are connected. Neurons in the primary field project to RM and R in both hemispheres and to AL, Pl, and the caudomedial (CM) field in the ipsilateral hemisphere. The primary fields in the two hemispheres are connected. Single injections into A1 and R often result in labeling of two or more columns of tissue in the ipsilateral and contralateral target fields. Cortico-cortical axon terminations are concentrated in layer IV of fields AL and RM and in upper layer III and layer IV of R and CM. In A1, axon terminals of neurons whose cell bodies lie in A1 in the opposite hemisphere are concentrated in upper layer III and layer IV; axon terminals of neurons located in field R of the same hemispheres are concentrated in layers I and II. Layer IV of Pl contains the greatest concentration of cortico-cortical axon terminals; the supragranular layers contain a somewhat lower concentration. Neurons in R project contralaterally in the anterior commissure while A1 neurons send their axons contralaterally in the corpus callosum.

Topography, cytoarchitecture, and sulcal patterns in primary somatic sensory cortex (SmI) prosimian primate, Perodicticus potto.

The topographic organization of the primary somatic sensory projection area (SmI) in relation to cytoarchitectural fields and sulcal patterns was examined in the prosimian primate Perodicticus potto. The area of cortex responding to low threshold (LT) cutaneous stimulation of the glabrous and hairy surfaces of the hand was determined by microelectrode mapping techniques, with standardized threshold stimuli for defining receptive fields. A single somatotopic projection of the two hand surfaces was found; the glabrous projection area is rostral to that of the hairy hand. Within both the glabrous and hairy areas, receptive fields on the distal digits are found anterior to those on the proximal hand. The glabrous hand projection area is coextensive with a dense granular area typical of koniocortex. The hairy hand area corresponds to a cytoarchitectural field which is less granular than the glabrous field. While koniocortex occupies the crown of the gyrus caudal to the coronally oriented sulcus, a large more rostral field, which contains both granule and large pyramidal cells, occupies the whole of the caudal bank of the sulcus. Force thresholds of many receptive fields (RFs) in Perodicticus were high both on the borders and within the LT area (perhaps because of the advanced age of these animals). However, the receptive field sizes for both the glabrous and hairy hand areas were of the same magnitude as those of Nycticebus (Carlson and FitzPatrick, '82). From the combined studies of three species of Lorisidae, Perodicticus, Galago (Carlson and Welt, '80), and Nycticebus (Carlson and FitzPatrick, '81), using similar mapping and stimulation techniques, both general and specific features of SmI hand area organization can be illustrated. A single projection of the glabrous and hairy hand is common to Perodicticus and Galago, but two glabrous projection areas are seen in Nycticebus. The projection area for the hand in Perodicticus is twice as large (relative to brain size) as in Galago and Perodicticus. The possible behavioral significance of increased differentiation of the hand area in Nycticebus and elaboration of the area in Perodicticus could be examined by study of hand use and tactile capacity in these same species.

Quality assurance in analysis of cryogenically stored liver tissue specimens from the NIST National Biomonitoring Specimen Bank (NBSB).

Portions of liver tissue specimens originally stored in the National Biomonitoring Specimen Bank (NBSB) and analyzed between 1980 and 1987 were re-analyzed in 1997 using instrumental neutron activation analysis (INAA) for the determination of 17 trace elements. Duplicate portions of each specimen had been stored at two different temperatures. The first was stored in a liquid nitrogen vapor-cooled freezer at -150 degrees C, standard NBSB storage conditions, and the other in an electric freezer maintained at -80 degrees C. Two portions of seven livers from each storage temperature were re-analyzed for this work. Results showed no changes in trace element content as a function of storage temperature, within the uncertainty of the method used. Results from these analyses agreed with results of initial analyses for most analytes in most sub-samples. Of the exceptions, five were due, in part, to an incorrect basis mass for the initial sub-specimen of one tissue, five with variable Zn results were attributed to difficulties in peak fitting for this element during INAA data processing, and the remaining were isolated differences discussed in this paper. Results of this work indicate that specimen storage and processing protocols are adequate to prevent noticeable contamination of specimens with trace elements, with the exception of Cr. Variability in Cr content was observed for the liver tissues which may have been caused by Cr contamination of the samples by the Teflon mill. Analyses of portions of Standard Reference Material (SRM) 1566a Oyster Tissue (certified in 1989) and SRM 1577a Bovine Liver (certified in 1982) were also included in this study for the purpose of quality control and to assess the stability of these freeze-dried powders that were stored at room temperature. No changes were observed in these materials.

Multiple strains of Coxiella burnetii are present in the environment of St. Paul Island, Alaska.

In 2010, Coxiella burnetii was identified at a high prevalence in the placentas of Northern fur seals (Callorhinus ursinus) collected at a single rookery on St. Paul Island Alaska; an area of the United States where the agent was not known to be present. As contamination was hypothesized as a potential cause of false positives, but nothing was known about environmental C. burnetii in the region, an environmental survey was conducted to look for the prevalence and distribution of the organism on the island. While environmental prevalence was low, two strains of the organism were identified using PCR targeting the COM1 and IS1111 genes. The two strains are consistent with the organism that has been increasingly identified in marine mammals as well as a strain type more commonly found in terrestrial environments and associated with disease in humans and terrestrial animals. Further work is needed to elucidate information regarding the ecology of this organism in this region, particularly in association with the coastal environment.

A genetic and molecular profile of third chromosome centric heterochromatin in Drosophila melanogaster.

In this review, we combine the results of our published and unpublished work with the published results of other laboratories to provide an updated map of the centromeric heterochromatin of chromosome 3 in Drosophila melanogaster. To date, we can identify more than 20 genes (defined DNA sequences with well-characterized functions and (or) defined genetic complementation groups), including at least 16 essential loci. With the ongoing emergence of data from genetic, cytological, and genome sequencing studies, we anticipate continued, substantial progress towards understanding the function, structure, and evolution of centric heterochromatin.

Essential genes in autosomal heterochromatin of Drosophila melanogaster.

We are taking two approaches to understanding the structure, function and regulation of essential genes within Drosophila heterochromatin. In the first, we have undertaken a genetic and molecular characterization of essential genes within proximal 3L heterochromatin. The expression of such 'resident' genes within a heterochromatic environment is paradoxical and poorly understood, given that the same environment can inactivate euchromatic sequences (position effect variegation, or PEV). A second approach involves the study of the local chromosomal environment of heterochromatic (het) genes, as assayed both biochemically, and via the effects of genetic modifiers of PEV, the latter being putative components important for het gene expression. Our results to date suggest that the three most proximal genes in 3L heterochromatin have key roles in development, and indicate strong effects of combinations of genetic modifiers of PEV on het gene expression.

The light gene of Drosophila melanogaster encodes a homologue of VPS41, a yeast gene involved in cellular-protein trafficking.

Mutations in a number of genes affect eye colour in Drosophila melanogaster; some of these "eye-colour" genes have been shown to be involved in various aspects of cellular transport processes. In addition, combinations of viable mutant alleles of some of these genes, such as carnation (car) combined with either light (lt) or deep-orange (dor) mutants, show lethal interactions. Recently, dor was shown to be homologous to the yeast gene PEP3 (VPS18), which is known to be involved in intracellular trafficking. We have undertaken to extend our earlier work on the lt gene, in order to examine in more detail its expression pattern and to characterize its gene product via sequencing of a cloned cDNA. The gene appears to be expressed at relatively high levels in all stages and tissues examined, and shows strong homology to VPS41, a gene involved in cellular-protein trafficking in yeast and higher eukaryotes. Further genetic experiments also point to a role for lt in transport processes: we describe lethal interactions between viable alleles of lt and dor, as well as phenotypic interactions (reductions in eye pigment) between allels of lt and another eye-colour gene, garnet (g), whose gene product has close homology to a subunit of the human adaptor complex, AP-3.

Essential genes in proximal 3L heterochromatin of Drosophila melanogaster.

We have further characterized essential loci within the centric heterochromatin of the left arm of chromosome 3 (3L) of Drosophila melanogaster, using EMS, radiation and P element mutagenesis. We failed to find any new essential genes, a result that suggests a lower-than-average gene density in this region. Mutations affecting expression of the most proximal gene [lethal 1, l1 or l(3)80Fj] act as dominant suppressors of Polycomb (Pc), behavior which is consistent with a putative trithorax group (trx-G) gene. The third gene to the left of the centromere [lethal 3, l3 or l(3)80Fh] is likely to correspond to verthandi (vtd), a known trx-G gene that plays a role in the regulation of hedgehog (hh) expression and signalling. The intervening gene [lethal 2, l2 or l(3)80Fi] is required throughout development, and mutant alleles have interesting phenotypes; in various allelic combinations that survive, we observe fertility, bristle, wing, eye and cuticle defects.