Ciliary neurotrophic factor activates leptin-like pathways and reduces body fat, without cachexia or rebound weight gain, even in leptin-resistant obesity

Ciliary Neurotrophic Factor (CNTF) was first characterized as a trophic factor for motor neurons in the ciliary ganglion and spinal cord, leading to its evaluation in humans suffering from motor neuron disease. In these trials, CNTF caused unexpected and substantial weight loss, raising concerns that it might produce cachectic-like effects. Countering this possibility was the suggestion that CNTF was working via a leptin-like mechanism to cause weight loss, based on the findings that CNTF acts via receptors that are not only related to leptin receptors, but also similarly distributed within hypothalamic nuclei involved in feeding. However, although CNTF mimics the ability of leptin to cause fat loss in mice that are obese because of genetic deficiency of leptin (ob/ob mice), CNTF is also effective in diet-induced obesity models that are more representative of human obesity, and which are resistant to leptin. This discordance again raised the possibility that CNTF might be acting via nonleptin pathways, perhaps more analogous to those activated by cachectic cytokines. Arguing strongly against this possibility, we now show that CNTF can activate hypothalamic leptin-like pathways in diet-induced obesity models unresponsive to leptin, that CNTF improves prediabetic parameters in these models, and that CNTF acts very differently than the prototypical cachectic cytokine, IL-1. Further analyses of hypothalamic signaling reveals that CNTF can suppress food intake without triggering hunger signals or associated stress responses that are otherwise associated with food deprivation; thus, unlike forced dieting, cessation of CNTF treatment does not result in binge overeating and immediate rebound weight gain.

Even-odd alternation in mass spectrum of thymine and uracil clusters: Evidence of intracluster photodimerization

Multiphoton ionization of thymine and uracil clusters generated by a supersonic molecular beam gave rise to a remarkable alternation of mass spectral intensities between even- and odd-numbered clusters. Such alternation was observed in clusters of up to 30 molecules. Excitation to the two lowest electronically excited states seemed to be a strong prerequisite. In view of the well known photodimerization reaction of thymine and uracil in the bulk phase, it is proposed that such alternation in the mass spectral intensity resulted from formation of photodimer units within the cluster on intense UV irradiation. Several analogues of thymine with no known propensity for photodimerization in the bulk phase did not exhibit any sign of such alternation in the cluster mass spectrum. The intrinsic UV window for photodimerization, and hence photoinduced mammalian mutagenesis, was estimated to be approximately 210–280 nm, significantly narrower than the previously reported bulk values of 150–300 nm.

Effects of Xylem pH on Transpiration from Wild-Type and flacca Tomato Leaves1 : A Vital Role for Abscisic Acid in Preventing Excessive Water Loss Even from Well-Watered Plants

The pH of xylem sap from tomato (Lycopersicon esculentum) plants increased from pH 5.0 to 8.0 as the soil dried. Detached wild-type but not flacca leaves exhibited reduced transpiration rates when the artificial xylem sap (AS) pH was increased. When a well-watered concentration of abscisic acid (0.03 μm) was provided in the AS, the wild-type transpirational response to pH was restored to flacca leaves. Transpiration from flacca but not from wild-type leaves actually increased in some cases when the pH of the AS was increased from 6.75 to 7.75, demonstrating an absolute requirement for abscisic acid in preventing stomatal opening and excessive water loss from plants growing in many different environments.

Formation of Holliday junctions by regression of nascent DNA in intermediates containing stalled replication forks: RecG stimulates regression even when the DNA is negatively supercoiled

Replication forks formed at bacterial origins often encounter template roadblocks in the form of DNA adducts and frozen protein–DNA complexes, leading to replication-fork stalling and inactivation. Subsequent correction of the corrupting template lesion and origin-independent assembly of a new replisome therefore are required for survival of the bacterium. A number of models for replication-fork restart under these conditions posit that nascent strand regression at the stalled fork generates a Holliday junction that is a substrate for subsequent processing by recombination and repair enzymes. We show here that early replication intermediates containing replication forks stalled in vitro by the accumulation of excess positive supercoils could be cleaved by the Holliday junction resolvases RusA and RuvC. Cleavage by RusA was inhibited by the presence of RuvA and was stimulated by RecG, confirming the presence of Holliday junctions in the replication intermediate and supporting the previous proposal that RecG could catalyze nascent strand regression at stalled replication forks. Furthermore, RecG promoted Holliday junction formation when replication intermediates in which the replisome had been inactivated were negatively supercoiled, suggesting that under intracellular conditions, the action of RecG, or helicases with similar activities, is necessary for the catalysis of nascent strand regression.

TAT peptide on the surface of liposomes affords their efficient intracellular delivery even at low temperature and in the presence of metabolic inhibitors

To achieve an efficient intracellular drug and DNA delivery, attempts were made to target microparticulate drug carriers into cytoplasm bypassing the endocytotic pathway. TAT peptides derived from the HIV-1 TAT protein facilitate intracellular delivery of proteins and small colloidal particles. We demonstrated that relatively large drug carriers, such as 200-nm liposomes, can also be delivered into cells by TAT peptide attached to the liposome surface. Liposomes were fluorescently labeled with membranotropic rhodamine-phosphatidylethanolamine or by entrapping FITC-dextran. Incubation of fluorescent TAT liposomes with mouse Lewis lung carcinoma cells, human breast tumor BT20 cells, and rat cardiac myocyte H9C2 results in intracellular localization of certain liposomes. Steric hindrances for TAT peptide⋅cell interaction (attachment of TAT directly to the liposome surface without spacer or the presence of a high MW polyethylene glycol on the liposome surface) abolish liposome internalization, evidencing the importance of direct contact of TAT peptide with the cell surface. Low temperature or metabolic inhibitors, sodium azide or iodoacetamide, have little influence on the translocation of TAT liposomes into cells, confirming the energy-independent character of this process. The approach may have important implications for drug delivery directly into cell cytoplasm.

Loss of tramtrack gene activity results in ectopic R7 cell formation, even in a sina mutant background.

We have screened a collection of transposable-element-induced mutations for those which dominantly modify the extra R7 phenotype of a hypomorphic yan mutation. The members of one of the identified complementation groups correspond to disruptions of the tramtrack (ttk) gene. As heterozygotes, ttk alleles increase the percentage of R7 cells in yan mutant eyes. Just as yan mutations increase ectopic R7 cell formation, homozygous ttk mutant eye clones also contain supernumerary R7 cells. However, in contrast to yan, the formation of these cells in ttk mutant eye tissue is not necessarily dependent on the activity of the sina gene. Furthermore, although yan mutations dominantly interact with mutations in the Ras1, Draf, Dsor1, and rolled (rl) genes to influence R7 cell development, ttk mutations only interact with yan and rl gene mutations to affect this signaling pathway. Our data suggest that yan and ttk both function to repress inappropriate R7 cell development but that their mechanisms of action differ. In particular, TTK activity appears to be autonomously required to regulate a sina-independent mechanism of R7 determination.