Assignment of human uroporphyrinogen decarboxylase (URO-D) to the p34 band of chromosome 1

Summary A cDNA probe corresponding to mRNA encoding human uroporphyrinogen decarboxylase (URO-D) was used to determine the chromosomal localization of the URO-D gene in the human genome. In agreement with previous studies, we have found that the locus for URO-D is located on chromosome 1 in hybrid cell mapping panels. The use of in sity hybridization allowed us to map the URO-D locus to band 1p34.Part of this work was presented as an abstract entitled Localization of the uroporphyrinogen decarboxylase gene to 1p34 band, by in situ hybridization, by M. G. Mattei, A. Dubart, D. Beaupain, M. Goossens, and J. F. Mattei, for a poster presentation at the 8th International Conference on Human Gene Mapping, Helsinki, August 4–10, 1985     

Depressed cytotoxic T-cell responses in previously treated Hodgkin's and non-Hodgkin's lymphoma patients

Summary Peripheral blood lymphocytes from 62 previously treated Hodgkin's and non-Hodgkin's lymphoma patients were tested for their ability to generate cytotoxic T lymphocytes in response to stimulation with allogeneic cells in mixed leukocyte culture. In most patients, including some in long-term unmaintained remission, extremely low cytotoxic responses were generated. To test whether these patients have circulating cells that suppress autologous lymphocytes from responding to alloantigens, patients' responding cells were passaged over columns of sepharose beads conjugated with histamine-rabbit serum albumin (Hist-RSA). This procedure has been shown to remove mouse suppressor cells and Concanavalin A(ConA)-induced human suppressor cells. Passage of patients' cells, prior to allogeneic stimulation, over columns of sepharose beads conjugated with Hist-RSA but not over control RSA columns, resulted in the isolation of lymphocytes that generated increased cytotoxic responses to alloantigens in 18 of 22 patients with initially low cytotoxic responses. These results suggest that the impaired ability of treated Hodgkin's and non-Hodgkin's lymphoma patients' lymphocytes to differentiate into cytotoxic T lymphocytes is at least in part due to the presence of circulating suppressor cells that bear histamine receptors.Scholar of the Leukemia Society of America     

Cytokinesis and Midzone Microtubule Organization in Caenorhabditis elegans Require the Kinesin-like Protein ZEN-4

Members of the MKLP1 subfamily of kinesin motor proteins localize to the equatorial region of the spindle midzone and are capable of bundling antiparallel microtubules in vitro. Despite these intriguing characteristics, it is unclear what role these kinesins play in dividing cells, particularly within the context of a developing embryo. Here, we report the identification of a null allele of zen-4, an MKLP1 homologue in the nematode Caenorhabditis elegans, and demonstrate that ZEN-4 is essential for cytokinesis. Embryos deprived of ZEN-4 form multinucleate single-celled embryos as they continue to cycle through mitosis but fail to complete cell division. Initiation of the cytokinetic furrow occurs at the normal time and place, but furrow propagation halts prematurely. Time-lapse recordings and microtubule staining reveal that the cytokinesis defect is preceded by the dissociation of the midzone microtubules. We show that ZEN-4 protein localizes to the spindle midzone during anaphase and persists at the midbody region throughout cytokinesis. We propose that ZEN-4 directly cross-links the midzone microtubules and suggest that these microtubules are required for the completion of cytokinesis.     

Soil respiration within riparian buffers and adjacent crop fields

We quantified rates of soil respiration among sites within an agricultural landscape in central Iowa, USA. The study was conducted in riparian cool-season grass buffers, in re-established multispecies (switchgrass + poplar) riparian buffers and in adjacent crop (maize and soybean) fields. The objectives were to determine the variability in soil respiration among buffer types and crop fields within a riparian landscape, and to identify those factors correlating with the observed differences. Soil respiration was measured approximately monthly over a two-year period using the soda-lime technique. Mean daily soil respiration across all treatments ranged from 0.14 to 8.3 g C m^–2 d^–1. There were no significant differences between cool-season grass buffers and re-established forest buffers, but respiration rates beneath switchgrass were significantly lower than those beneath cool-season grass. Soil respiration was significantly greater in both buffer systems than in the cropped fields. Seasonal changes in soil respiration were strongly related to temperature changes. Over all sites, soil temperature and soil moisture together accounted for 69% of the seasonal variability in soil respiration. Annual soil respiration rates correlated strongly with soil organic carbon (R = 0.75, P < 0.001) and fine root (<2 mm) biomass (R = 0.85, P < 0.001). Annual soil respiration rates averaged 1140 g C m^–2 for poplar, 1185 g C m^–2 for cool-season grass, 1020 g C m^–2 for switchgrass, 750 g C m^–2 for soybean and 740 g C m^–2 for corn. Overall, vegetated buffers had significantly higher soil respiration rates than did adjacent crop fields, indicating greater soil biological activity within the buffers.     

Rapid epithelial-sheet sealing in the Caenorhabditis elegans embryo requires cadherin-dependent filopodial priming.

BACKGROUND: During embryonic development, epithelia with free edges must join together to create continuous tissues that seal the interior of the organism from the outside environment; failure of epithelial sealing underlies several common human birth defects. Sealing of epithelial sheets in embryos can be extremely rapid, dramatically exceeding the rate of adherens junction formation by epithelial cells in culture or during healing of epithelial wounds. Little is known about the dynamic redistribution of cellular junctional components during such events in living embryos. RESULTS: We have used time-lapse, multiphoton laser-scanning microscopy and green fluorescent protein fusion proteins to analyze the sealing of the Caenorhabditis elegans epidermis in living embryos. Rapid recruitment of alpha-catenin to sites of filopodial contact between contralateral migrating epithelial cells, concomitant with clearing of cytoplasmic alpha-catenin, resulted in formation of nascent junctions; this preceded the formation of mature junctions. Surprisingly, upon inactivation of the entire cadherin-catenin complex, only adhesive strengthening between filopodia was reproducibly affected. Other ventral epidermal cells, which did not extend filopodia and appeared to seal along the ventral midline by coordinated changes in cell shape, successfully adhered in the absence of these proteins. CONCLUSIONS: We propose that 'filopodial priming' - prealignment of bundled actin in filopodia combined with the rapid recruitment of alpha-catenin from cytoplasmic reserves at sites of filopodial contact - accounts for the rapid rate of sealing of the embryonic epidermis of C. elegans. Filopodial priming may provide a general mechanism for rapid creation of adherens junctions during epithelial-sheet sealing in embryos.     

High-density hapten labeling and HRP conjugation of oligonucleotides for use as in situ hybridization probes to detect mRNA targets in cells and tissues.

Oligonucleotides that carry a detectable label can be used to probe for mRNA targets in in situ hybridization experiments. Oligonucleotide probes (OPs) have several advantages over cDNA probes and riboprobes. These include the easy synthesis of large quantities of probe, superior penetration of probe into cells and tissues, and the ability to design gene- or allele-specific probes. One significant disadvantage of OPs is poor sensitivity, in part due to the constraints of adding and subsequently detecting multiple labels per oligonucleotide. In this study, we compared OPs labeled with multiple detectable haptens (such as biotin, digoxigenin, or fluorescein) to those directly conjugated with horseradish peroxidase (HRP). We used branching phosphoramidites to add from two to 64 haptens per OP and show that in cells, 16-32 haptens per OP give the best detection sensitivity for mRNA targets. OPs were also made by directly conjugating the same oligonucleotide sequences to HRP. In general, the HRP-conjugated OPs were more sensitive than the multihapten versions of the same sequence. Both probe designs work well both on cells and on formaldehyde-fixed, paraffin-embedded tissues. We also show that a cocktail of OPs further increases sensitivity and that OPs can be designed to detect specific members of a gene family. This work demonstrates that multihapten-labeled and HRP-conjugated OPs are sensitive and specific and can make superior in situ hybridization probes for both research and diagnostic applications.     

Soil Temperature, Not Aboveground Plant Productivity, Best Predicts Intra-Annual Variations of Soil Respiration in Central Iowa Grasslands

Soil respiration (R_SOIL) is the second largest carbon flux between terrestrial systems and the atmosphere, with a magnitude 10 times greater than anthropogenic carbon dioxide production. Therefore, it is important that we understand, and be able to predict, how R_SOIL responds to climate change. Although a positive, significant temperature effect on R_SOIL has long been recognized, recent studies emphasize the overriding importance of current photosynthesis in controlling R_SOIL. We tested the hypothesis that model inclusion of intra-annual variations in aboveground net primary productivity (ANPP) significantly improves R_SOIL estimates over predictions based on soil temperature alone. We also evaluated the possibility that canopy production is less directly linked to R_SOIL, by testing the hypothesis that intersite differences in R_SOIL correlate more strongly with root biomass than with ANPP. We tested these hypotheses by measuring R_SOIL, ANPP, and root biomass at four Iowa grasslands that differed in aboveground growth phenology and productivity. Among all sites, intra-annual variations in R_SOIL were most strongly related to soil temperature (R ² = 0.89), not ANPP (R ² = 0.53). All sites responded identically to changes in soil temperature (site-by-temperature P = 0.53), but inconsistently to variation in aboveground dynamics (site-by-canopy P < 0.0001). Incorporating canopy dynamics into temperature-based predictive models improved model R ² by a maximum of 0.01. Among-site differences in R_SOIL were related to root biomass (P < 0.001) but not ANPP (P = 0.34). We found no useful linkage between canopy characteristics and intra-annual or site-specific R_SOIL predictions, perhaps because shoot and root dynamics were not consistently linked through time or among sites.