Analysis of the pattern of gene expression during human adipogenesis by DNA microarray

High density DNA microarrays containing over 5000 cDNA clones were used to carry out a comprehensive investigation of gene expression during adipogenesis. Complex probes synthesized from total RNA were hybridized to the arrays to determine the level of mRNA expression of each arrayed gene. Thirty three genes (29 known and 4 ESTs with no identified homologies) have been found to alter their level of expression more than 2.5-fold after differentiation. The quantitative measurement by DNA array was in good agreement with conventional Northern blot analysis of selected genes. Our results demonstrate that utilization of a DNA array is a speedy, efficient and quantitative approach to profile the expression of a large number of genes.     

Measurements of phosphorus uptake by macrophytes and epiphytes from the LaPlatte River (VT) using 32P in stream microcosms

1. Phosphorus (P) uptake by macrophytes and epiphytes from the LaPlatte River (VT) was examined in the laboratory by adding ³²PO₄‐P to recirculating stream microcosms.
2. Water, plugs of sediment and plants were removed from the river and placed into the microcosms. ³²PO₄‐P was then added either to the water or the sediment, and its incorporation into plants and epiphytes was monitored over 3 days. Uptake was examined at both ambient (5 μg L^–1) and increased (50 μg L^–1) soluble reactive phosphorus (SRP) concentrations. A computer program was developed to fit curves to the radiotracer data and calculate rate constants for the simultaneous transfer of ³²P among compartments.
3. Both macrophytes and epiphytes removed P from the water, but epiphyte uptake of P was more rapid. Phosphate enrichment stimulated P uptake by both macrophytes and epiphytes. Macrophytes also obtained P from the sediment. The relative contribution of P to macrophytes from the water vs. that from the sediment appeared to vary with SRP in the overlying water. Accurate estimates of rates of P uptake from sediments by macrophytes were difficult to obtain however, due to very low and highly variable unit rate constants for P uptake and uncertainty about the magnitude of the phosphate pool available for uptake.
4. SRP concentrations were greater in the overlying water than in the sediment pore water of stream microcosms in the present study. Numerous reports in the literature have suggested that this condition favours uptake by macrophyte stems and leaves rather than by roots.
5. Phosphate uptake from the water by macrophytes in shallow streams may be more common than for macrophytes in lakes.     

Differential expression of genes encoding TGFs beta 1, beta 2, and beta 3 during murine palate formation.

Transforming growth factor beta 1 (TGF beta 1) has been shown to have multiple effects on primary cultures of palate-derived cell types. We report the analysis, by in situ hybridization, of RNA expression for three different TGF beta isoforms (TGF beta 1, beta 2, and beta 3) during murine embryonic palate development. Differential expression of the three TGF beta genes is seen in the palatal shelves in mesenchymal and epithelial cells known to be involved in the morphogenesis of this organ. Taken together, these results suggest that the TGF beta s act as endogenous factors involved in the formation of the mammalian palate.     

Segmental identity and cerebellar granule cell induction in rhombomere 1

Background  

Cerebellar granule cell precursors are specifically generated within the hindbrain segment, rhombomere 1, which is bounded rostrally by the midbrain/hindbrain isthmus and caudally by the boundary of the Hoxa2 expression domain. While graded signals from the isthmus have a demonstrable patterning role within this region, the significance of segmental identity for neuronal specification within rhombomere 1 is unexplored. We examined the response of granule cell precursors to the overexpression of Hoxa2, which normally determines patterns of development specific to the hindbrain. How much does the development of the cerebellum, a midbrain/hindbrain structure, reflect its neuromeric origin as a hindbrain segment?     

Spectroscopic methods for analysis of protein secondary structure

Several methods for determination of the secondary structure of proteins by spectroscopic measurements are reviewed. Circular dichroism (CD) spectroscopy provides rapid determinations of protein secondary structure with dilute solutions and a way to rapidly assess conformational changes resulting from addition of ligands. Both CD and Raman spectroscopies are particularly useful for measurements over a range of temperatures. Infrared (IR) and Raman spectroscopy require only small volumes of protein solution. The frequencies of amide bands are analyzed to determine the distribution of secondary structures in proteins. NMR chemical shifts may also be used to determine the positions of secondary structure within the primary sequence of a protein. However, the chemical shifts must first be assigned to particular residues, making the technique considerably slower than the optical methods. These data, together with sophisticated molecular modeling techniques, allow for refinement of protein structural models as well as rapid assessment of conformational changes resulting from ligand binding or macromolecular interactions. A selected number of examples are given to illustrate the power of the techniques in applications of biological interest.     

Sequence-specific 1H NMR assignment and secondary structure of neuropeptide Y in aqueous solution

Sequence-specific assignment of the 1H NMR spectrum of the 36 amino acid polypeptide porcine neuropeptide Y (pNPY) at pH 3.1 is reported. It was achieved by use of standard two-dimensional techniques and by a combination of the sequential and main-chain-directed assignment strategies. The secondary structure was derived from inspection of the nuclear Overhauser spectra, slow hydrogen-deuterium exchange effects, chemical shifts of main-chain HA resonances, and coupling constants. These studies indicate that the C-terminal segment (residues 11-36) folds into an amphiphilic alpha-helix; the N-terminal segment, containing three prolines in both cis and trans conformations, assumes no regular structure. CD studies of pNPY at pH 3.1 and 7.4 show an increase in ordered structure at neutral pH. The difference spectrum, however, is typical of an alpha-helix and suggests a stabilization of residues 11-36, possibly via Maxfield-Scheraga pair interactions involving side-chain residues. This is supported by a comparison of the one-dimensional 1H NMR spectra of pNPY at pH 3.1 and 7.4, where no remarkable differences are observed.