. The functional relevance of the interaction between IL1RAPL1 and NCS-1 was also suggested by the reduction of neurite elongation observed in nerve growth factor (NGF)-treated IL1RAPL1 cells, a phenotype rescued by NCS-1 inactivation. Because both proteins are highly expressed in neurons, these results suggest that IL1RAPL1-related mental retardation could result from a disruption of N-VGCC and/or NCS-1-dependent synaptic and neuronal activities.PC12 cells ͉ neuronal calcium sensor-1 ͉ X-linked mental retardation ͉ exocytosis C ognitive impairment or mental retardation (MR) affects Ϸ2% of the population, leading to moderate (IQ Ͻ70) up to severe (IQ Ͻ50) handicap. The underlying causes are extremely heterogeneous, including genetic causes, many of which are X-linked (XLMR) (1, 2). Null mutations in the IL1-receptor accessory protein-like 1 gene (IL1RAPL1) [National Center for Biotechnology Information (NCBI) accession no. NM 014271.2] have been involved in a nonsyndromic form of XLMR (3). IL1RAPL1 and the closely related protein IL1RAPL2 (NCBI accession no. NM 017416) belong to a previously unrecognized class of the interleukin-1/toll receptor family characterized by the presence of a 150-aa C terminus domain with unknown function (3, 4). IL1RAPL1 and IL1RAPL2 have specific, but not redundant, temporal and spatial expression pattern in the brain, Il1rapl1, the mouse homologue of IL1RAPL1, being specifically expressed in adult brain structures that are known to be involved in the hippocampal memory system (3).Most of physiological and biological properties of IL1RAPL proteins remain largely unknown. Recent studies have demonstrated that expression of IL1RAPL1 in PC12 cells leads to a profound inhibition of ATP-induced growth hormone (GH) release (4). The underlying cellular pathways remain to be elucidated; however, IL1RAPL1 has been shown to interact by way of its 150-aa C-terminal domain with the neuronal calcium sensor-1 protein (NCS-1), a protein widely expressed in neurons (5) and the related chromaffin and PC12 cells (6).NCS-1 belongs to a large Ca 2ϩ -binding protein family (7) implicated in the regulation of Ca 2ϩ -dependent exocytosis (6) through its activation of PI 4 kinase and PIP 2 formation (8-10). Moreover, NCS-1 modulates Ca 2ϩ channels trafficking and activity in various cellular models (11, 12) and has effects on synaptic transmission by way of activity-dependent facilitation of P/Q-type calcium currents at presynaptic nerve terminals (13). Finally, at the behavioral level, NCS-1 appears also to be involved in associative learning and memory (14).Here, we focus on IL1RAPL1 functions by studying the physiological impact of IL1RAPL1/NCS-1 interaction in PC12 cells, a well known model of both nerve growth factor (NGF)-induced neuronal differentiation and Ca 2ϩ -dependent exocytosis. We show that expression of IL1RAPL1 in PC12 cells, which do not normally express the IL1RAPL1 protein, mediates, through IL1RAPL1/NSC-1 interaction, a down-regulation of N-type voltage-gated calcium channel (N-VGCC) ac...

The rat aldolase C gene encodes a glycolytic enzyme strongly expressed in adult brain. We previously reported that a 115-base pair (bp) promoter fragment was able to ensure the brain-specific expression of the chloramphenicol acetyltransferase (CAT) reporter gene in transgenic mice, but only at a low level (Thomas, M., Makeh, I., Briand, P., Kahn, A., and Skala, H. (1993) Eur. J. Biochem. 218, 143-151). Here we show that in vivo activation of this promoter at a high level requires cooperation between an upstream 0.6-kilobase pair (kb) fragment and far upstream sequences. In the 0.6-kb region, a 28-bp DNA element is shown to include overlapping in vitro binding sites for POU domain regulatory proteins and for the Winged Helix hepatocyte nuclear factor-3␤ factor. An hepatocyte nuclear factor-3␤-binding site previously described in the short proximal promoter fragment is also shown to interact in vitro with POU proteins, although with a lower affinity than the 28-bp motif. Additional binding sites for POU factors were detected in the upstream 0.6-kb sequences. Progressive deletion in this region resulted in decreased expression levels of the transgenes in mice, suggesting synergistic interactions between these multiple POUbinding sites. We propose that DNA elements characterized by a dual binding specificity for both POU domain and Winged Helix transcription factors could play an essential role in the brain-specific expression of the aldolase C gene and other neuronal genes.

A rat brain cDNA library was screened at low stringency with an aldolase B cDNA probe corresponding to the coding sequence of the mRNA, then at high stringency with a 3' non-coding aldolase A cDNA probe. One clone, which hybridized only under the first conditions, was further characterized and used to screen the library again.Two overlapping clones, complementary to aldolase C mRNA, were obtained. They cover the 113 cdrboxyterminal coding residues and the 3' non-coding region up to the poly(A) tail. Their nucleotide sequence was determined. In the coding region the overall homology with aldolase A was 67% at the nucleotide level and 76% at the protein level. With aldolase B these values were 63% and 65% respectively. The 3' non-coding region was 380 bases long and did not exhibit any homology with the untranslated 3' extension of aldolase A and B mRNAs.Southern blot analysis indicates that probably a single aldolase C gene exists per haploid genome. Aldolase C mRNA was detected at low concentration in practically all the foetal tissues and its expression markedly and rapidly decreased after birth. In brain the concentration of aldolase C mRNA remained high and stable even after birth. Aldolase C mRNA is approximately 50-fold more abundant in brain than in foetal tissues, which are the richest in messenger RNA.In the course of azo-dye hepatocarcinogenesis the aldolase C gene is re-expressed early, with a maximum at the 4th week of carcinogenic diet, which probably corresponds to the maximal proliferation of the oval cells.