SINGLE-CELL PIGMENTATION OF PORPHYRA LINEARIS ANALYZED BY FOURIER TRANSFORM MULTI-PIXEL SPECTROSCOPY AND IMAGE ANALYSIS1

The novel method of Fourier transform multi-pixel spectroscopy was used for the nondestructive analysis of and comparison of pigmentation in different regions of live thalli of the red alga Porphyra linearis. Because the thallus in this alga consists of a monolayer of nonoverlapping cells, we were able to analyze the pigmentation of single cells by combining light absorbance with natural fluorescence data. From the image of each cell in the vegetative male and female reproductive and holdfast regions, more than 4 ± 10⁴ fluorescence and absorbance spectra were obtained. Specific pigments in the different regions were localized by the use of a software program of similarity mapping followed by image construction. The reconstructed images revealed subcellular localization of each pigment according to specific spectroscopic fingerprints. The results showed that the vegetative and female reproductive cell types had a significantly higher content of phycoerythrin than of phycocyanin, and quite similar chlorophyll a levels. Most of the holdfast cells were poorly pigmented, but had more chlorophyll a than phycoerythrin or phycocyanin. The male reproductive cells contained only traces of pigments. Thus, by using Fourier transform multipixel spectroscopy, we were able to characterize the pigmentation of different regions of the thallus and follow the distribution patterns of the different pigments on the subcellular level along the differentiation gradient of the alga.     

Spatial and Temporal Biogeography of Soil Microbial Communities in Arid and Semiarid Regions

Microbial communities in soils may change in accordance with distance, season, climate, soil texture and other environmental parameters. Microbial diversity patterns have been extensively surveyed in temperate regions, but few such studies attempted to address them with respect to spatial and temporal scales and their correlations to environmental factors, especially in arid ecosystems. In order to fill this gap on a regional scale, the molecular fingerprints and abundance of three taxonomic groups – Bacteria, α-Proteobacteria and Actinobacteria – were sampled from soils 0.5–100 km apart in arid, semi-arid, dry Mediterranean and shoreline Mediterranean regions in Israel. Additionally, on a local scale, the molecular fingerprints of three taxonomic groups – Bacteria, Archaea and Fungi – were sampled from soils 1 cm–500 m apart in the semi-arid region, in both summer and winter. Fingerprints of the Bacteria differentiated between all regions (P<0.02), while those of the α-Proteobacteria differentiated between some of the regions (0.01<P<0.09), and actinobacterial fingerprints were similar among all regions (P>0.05). Locally, fingerprints of archaea and fungi did not display distance-decay relationships (P>0.13), that is, the dissimilarity between communities did not increase with geographic distance. Neither was this phenomenon evident in bacterial samples in summer (P>0.24); in winter, however, differences between bacterial communities significantly increased as the geographic distances between them grew (P<0.01). Microbial community structures, as well as microbial abundance, were both significantly correlated to precipitation and soil characteristics: texture, organic matter and water content (R²>0.60, P<0.01). We conclude that on the whole, microbial biogeography in arid and semi-arid soils in Israel is determined more by specific environmental factors than geographic distances and spatial distribution patterns.     

Deterministic Somatic Cell Reprogramming Involves Continuous Transcriptional Changes Governed by Myc and Epigenetic-Driven Modules

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Axoplasmic importins enable retrograde injury signaling in lesioned nerve

Axoplasmic proteins containing nuclear localization signals (NLS) signal retrogradely by an unknown mechanism in injured nerve. Here we demonstrate that the importin/karyopherin alpha and beta families underlie this process. We show that importins are found in axons at significant distances from the cell body and that importin beta protein is increased after nerve lesion by local translation of axonal mRNA. This leads to formation of a high-affinity NLS binding complex that traffics retrogradely with the motor protein dynein. Trituration of synthetic NLS peptide at the injury site of axotomized dorsal root ganglion (DRG) neurons delays their regenerative outgrowth, and NLS introduction to sciatic nerve concomitantly with a crush injury suppresses the conditioning lesion induced transition from arborizing to elongating growth in L4/L5 DRG neurons. These data suggest a model whereby lesion-induced upregulation of axonal importin beta may enable retrograde transport of signals that modulate the regeneration of injured neurons.     

Children diagnosed with diabetes during infancy have unique clinical characteristics

OBJECTIVE: Characterizing clinical and biochemical features of children diagnosed with diabetes mellitus between the ages of 6-24 months. DESIGN AND METHODS: Medical records of 42 children diagnosed with diabetes mellitus at age of 6-24 months were reviewed for gender, ethnic origin, family medical history, clinical and biochemical features at onset of diabetes compared with 60 diabetic patients diagnosed at age 5-16 years. RESULTS: Children diagnosed at 6-24 months had at onset more symptoms of apathy, restlessness, hyperglycemia during acute illness and a lower rate of remission than those diagnosed at older age (p < 0.001), significantly more episodes of diabetic ketoacidosis (83% vs. 40%, p < 0.001), lower HbA1c levels (mean 11.6 +/- 3.4 vs. 13.75 +/- 3.4%, p < 0.05) and a higher rate of celiac disease (12% vs. 3%, p = 0.046). There were no significant differences as to other autoimmune diseases. CONCLUSIONS: Patients with diabetes presenting at 6-24 months might be associated with a different clinical pattern and higher rate of celiac disease than diabetes presenting later in life. Understanding the nature and course of diabetes in this age group is crucial for planning interventional and preventive programs.     

The Ras Antagonist,Farnesylthiosalicylic Acid (FTS), Decreases Fibrosis and Improves Muscle Strength in dy2J/dy2J Mouse Model of Muscular Dystrophy

The Ras superfamily of guanosine-triphosphate (GTP)-binding proteins regulates a diverse spectrum of intracellular processes involved in inflammation and fibrosis. Farnesythiosalicylic acid (FTS) is a unique and potent Ras inhibitor which decreased inflammation and fibrosis in experimentally induced liver cirrhosis and ameliorated inflammatory processes in systemic lupus erythematosus, neuritis and nephritis animal models. FTS effect on Ras expression and activity, muscle strength and fibrosis was evaluated in the dy^2J/dy^2J mouse model of merosin deficient congenital muscular dystrophy. The dy^2J/dy^2J mice had significantly increased RAS expression and activity compared with the wild type mice. FTS treatment significantly decreased RAS expression and activity. In addition, phosphorylation of ERK, a Ras downstream protein, was significantly decreased following FTS treatment in the dy^2J/dy^2J mice. Clinically, FTS treated mice showed significant improvement in hind limb muscle strength measured by electronic grip strength meter. Significant reduction of fibrosis was demonstrated in the treated group by quantitative Sirius Red staining and lower muscle collagen content. FTS effect was associated with significantly inhibition of both MMP-2 and MMP-9 activities. We conclude that active RAS inhibition by FTS was associated with attenuated fibrosis and improved muscle strength in the dy^2J/dy^2J mouse model of congenital muscular dystrophy.     

High-nitrogen compost as a medium for organic container-grown crops

Compost was tested as a medium for organic container-grown crops. Nitrogen (N) loss during composting of separated cow manure (SCM) was minimized using high C/N (wheat straw, WS; grape marc, GM) or a slightly acidic (orange peels, OP) additives. N conservation values in the resultant composts were 82%, 95% and 98% for GM-SCM, OP-SCM and WS-SCM, respectively. Physical characteristics of the composts were compatible with use as growing media. The nutritional contribution of the composts was assessed using cherry tomato (Lycopersicon esculantum Mill.) and by means of incubation experiments. Media were either unfertilized or fertilized with guano (sea-bird manure). Plant responses suggest that N availability is the main variable affecting growth. Unfertilized OP-SCM and WS-SCM supplied the N needed for at least 4 months of plant growth. Root-galling index (GI) of tomato roots and number of eggs of the nematode Meloidogyne javanica were reduced by the composts, with the highest reduction obtained by OP-SCM and WS-SCM, at 50% concentrations. These composts, but not peat, reduced the incidence of crown and root-rot disease in tomato as well as the population size of the causal pathogen, Fusarium oxysporum f. sp. radicis-lycopersici.     

Neurodegeneration induces upregulation of Beclin 1

Autophagy, a bulk degradation of subcellular constituents, is activated in normal cell growth and development, and represents the major pathway by which the cell maintains a balance between protein synthesis and protein degradation. Autophagy was documented in several neurodegenerative diseases, and under stress conditions the autophagic process can lead to cell death (type II programmed cell death). Beclin 1 is a Bcl-2 interacting protein that was previously found to promote autophagy. We have used Beclin 1 protein as a marker for autophagy following traumatic brain injury in mice. We demonstrated a dramatic elevation in Beclin 1 levels near the injury site. Interestingly Beclin 1 elevation starts at early stages post injury (4 h) in neurons and 3 days later in astrocytes. In both cell types it lasts for at least three weeks. Neuronal cells, but not astrocytes, that overexpress Beclin 1 may exhibit damaged DNA but without changes in nuclear morphology. These observations may indicate that not all the Beclin1 overexpressing cells will die. The elevation of Beclin 1 at the site of injury may represent enhanced autophagy as a mechanism to discard injured cells and reduce damage to cells by disposing of injured components.     

Protein kinase Calpha regulates insulin receptor signaling in skeletal muscle

Certain PKC isoforms are stimulated by insulin and interact with IR as well as with IRS, but it is still not clear if specific PKC isoforms regulate IR signaling directly or through IRS-1. PKCalpha may regulate IRS activity in response to insulin. We investigated the possibility that PKCalpha may be important in insulin signaling. Studies were conducted on skeletal muscle in adult mice and on L6 skeletal cells. PKCalpha is constitutively associated with IRS-1, and insulin stimulation of PKCalpha causes disassociation of the two proteins within 5 min. Blockade of PKCalpha inhibited insulin-induced disassociation of PKCalpha from IRS1. Selective inhibition of PKCalpha increased the ability of insulin to reduce blood glucose levels. Insulin stimulation activates PKB and increases the association of PKCalpha with PKB. Blockade of PKCalpha increased threonine phosphorylation of PKB. We suggest that PKCalpha regulates insulin signaling in skeletal muscle through its disassociation from IRS-1 and association with PKB.