Xenopus Xpat protein is a major component of germ plasm and may function in its organisation and positioning.

In many animals, including Drosophila, C. elegans, zebrafish and Xenopus, the germ line is specified by maternal determinants localised in a distinct cytoplasmic structure called the germ plasm. This is consists of dense granules, mitochondria, and specific localised RNAs. We have characterised the expression and properties of the protein encoded by Xpat, an RNA localised to the germ plasm of Xenopus. Immunofluorescence and immunoblotting showed that this novel protein is itself a major constituent of germ plasm throughout oogenesis and early development, although it is also present in other regions of oocytes and embryos, including their nuclei. We found that an Xpat-GFP fusion protein can localise correctly in cultured oocytes, in early oocytes to the 'mitochondrial cloud', from which germ plasm originates, and in later oocytes to the vegetal cortex. The localisation process was microtubule-dependent, while cortical anchoring required microfilaments. Xpat-GFP expressed in late stage oocytes assembled into circular fields of multi-particulate structures resembling endogenous fields of germ plasm islands. Furthermore these structures could be induced to form at ectopic sites by manipulation of culture conditions. Ectopic Xpat-GFP islands were able to recruit mitochondria, a major germ plasm component. These data suggest that Xpat protein has an important role in Xenopus germ plasm formation, positioning and maintenance.

Food deprivation differentially modulates orexin receptor expression and signaling in rat hypothalamus and adrenal cortex.

Although starvation-induced biochemical and metabolic changes are perceived by the hypothalamus, the adrenal gland plays a key role in the integration of metabolic activity and energy balance, implicating feeding as a major synchronizer of rhythms in the hypothalamic-pituitary-adrenal (HPA) axis. Given that orexins are involved in regulating food intake and activating the HPA axis, we hypothesized that food deprivation, an acute challenge to the systems that regulate energy balance, should elicit changes in orexin receptor signaling at the hypothalamic and adrenal levels. Food deprivation induced orexin type 1 (OX1R) and 2 (OX2R) receptors at mRNA and protein levels in the hypothalamus, in addition to a fivefold increase in prepro-orexin mRNA. Cleaved peptides OR-A and OR-B are also elevated at the protein level. Interestingly, adrenal OX1R and OX2R levels were significantly reduced in food-deprived animals, whereas there was no expression of prepro-orexin in the adrenal gland in either state. Food deprivation exerted a differential effect on OXR-G protein coupling. In the hypothalamus of food deprived rats compared with controls, a significant increase in coupling of orexin receptors to Gq, Gs, and Go was demonstrated, whereas coupling to Gi was relatively less. However, in the adrenal cortex of the food-deprived animal, there was decreased coupling of orexin receptors to Gs, Go, and Gq and increased coupling to Gi. Subsequent second-messenger studies (cAMP/IP3) have supported these findings. Our data indicate that food deprivation has differential effects on orexin receptor expression and their signaling characteristics at the hypothalamic and adrenocortical levels. These findings suggest orexins as potential metabolic regulators within the HPA axis both centrally and peripherally.