Identification of target mRNAs for the clock-controlled RNA-binding protein Chlamy 1 from Chlamydomonas reinhardtii.

An endogenous clock regulates the temporal expression of genes/mRNAs that are involved in the circadian output pathway. In the green alga Chlamydomonas reinhardtii, a clock-controlled RNA-binding protein (Chlamy 1) was identified recently, which represents an analog of the circadian trans-acting factor CCTR from the phylogenetically diverse alga Gonyaulax polyedra. In order to identify in C. reinhardtii target mRNAs that can be recognized by Chlamy 1, gel mobility-shift assays and UV-crosslinking experiments were carried out, and revealed that this protein interacts specifically with the 3' untranslated regions of several mRNAs and recognizes them all via a common cis-acting element, composed of at least seven UG repeats. By using competition assays, it was found that the affinity of Chlamy 1 is highest for mRNAs whose products are key components of nitrogen and CO2 metabolism. Since the activities of enzymes involved in nitrogen metabolism vary in a temporal pattern that is opposite in phase to that of Chlamy 1 binding activity, the protein may repress the translation of the cognate mRNAs.

Facilitative effects of an adenosine A1/A2 receptor blockade on spatial memory performance of rats: selective enhancement of reference memory retention during the light period.

The present experiment was designed to examine the role of adenosine in spatial working and reference memory in rats using an 8-arm radial maze task which requires the integrity of the hippocampal formation. We investigated the effects of the unselective adenosine A1/A2 receptor antagonist theophylline on acquisition and retention of spatial working and reference memory. As there is evidence that brain extracellular adenosine levels vary significantly during the light-dark cycle, we tested the effects of theophylline both during the light and the dark period. Acquisition of the task was investigated for 10 consecutive days after rats received daily injections of vehicle or theophylline (15 mg/kg, intraperitoneally). Retention was tested in two nondrug sessions 7 and 14 days after completion of acquisition. The results demonstrate that in saline-treated control rats acquisition and retention of reference memory and, to a lesser extent, working memory was superior in the dark period. The results further revealed that daily administration of theophylline interacted with days to selectively enhance reference memory acquisition in the light, but not in the dark, period. In addition, reference memory retention was significantly enhanced in those rats who learned the task under theophylline treatment during the light period. Overall, the results show that in saline-treated control rats the effectiveness of acquisition and retention of spatial information in a radial maze strongly depends on the time of day. The higher levels of maze performance in the dark period might be related to a better functioning of involved brain systems in the active period of the rat. Furthermore, theophylline-induced blockade of adenosine A1/A2 receptors in the light, but not in the dark, period selectively enhanced reference memory acquisition and retention. Variations of brain extracellular adenosine levels during the light-dark cycle might account for the restriction of reference memory enhancing effects of theophylline to the light period.