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We report that extracellular matrix and neurons modulate the expression of occludin, one of the main components of tight junctions, by rat brain endothelial cells (RBE4.B). Of the three extracellular matrix proteins which we tested (collagen I, collagen IV, and laminin), collagen IV stimulated at the best the expression of occludin mRNA. The corresponding protein, however, was not synthesized. Significant amounts of occludin accumulated only when RBE4.B cells were cultured on collagen IV-coated inserts, in the presence of cortical neurons, plated on laminin-coated companion wells. Finally, occludin segregated at the cell periphery, only when endothelial cells were co-cultured with neurons for > or = 1 week.

During maturation of mammalian brain, variants of both linker (i.e. H1°) and core (i.e. H3.3) histone proteins accumulate in nerve cells. As the concentration of the corresponding transcripts decreases, in postmitotic cells, even if the genes are actively transcribed, it is likely that regulation of variant histone expression has relevant post-transcriptional components and that cellular factors affect histone mRNA stability and/or translation. Here we report that PIPPin, a protein that is highly enriched in the rat brain and contains a coldshock domain, binds with high specificity to the transcripts that encode H1°and H3.3 histone variants. Both mRNAs are bound through the very end of their 3-untranslated region that encompasses the polyadenylation signal. Although PIPPin is present both in the cytoplasm and the nucleus of nerve cells, PIPPin-RNA complexes can be obtained only from nuclear extracts. The results of two-dimensional electrophoretic analysis suggest that a relevant proportion of nuclear PIPPin is more acidic than expected, thus suggesting that its RNA binding activity might be modulated by post-translational modifications, such as phosphorylation.During development of an organism and tissue differentiation, chromatin must be remodeled to permit entrance of transcription factors and hence expression of genes at the right places and times. Although a critical moment for setting new patterns of chromatin organization is the S phase of the cell cycle, it is now clear that chromatin can be remodeled also in the absence of DNA replication, by energy consuming complexes (1-4). The possibility that remodeling also allows entrance, at topologically defined regions of the nucleus, of specific histone isotypes, which might locally modify chromatin organization even more, is provocative and deserves of attention.We previously demonstrated that, in the developing rat brain, the concentration of H1°and H3.3 mRNAs decreases between the embryonal day 18 (E18) and the postnatal day 10 (P10), whereas the corresponding genes are transcribed at the same rate at any stage studied, suggesting that the two genes are regulated mainly at post-transcriptional level (5, 6). As post-transcriptional control processes, including regulation of splicing (7), vectorial transport of mature mRNAs (8 -10), regulation of mRNA stability (11-13), and availability for translation (14, 15), are mediated by several classes of RNA-binding proteins (for review, see , it is likely that developing rat brain contains mRNA-binding factors involved in the binding and regulation of mRNAs encoding histone variants. We reported in a previous paper (19) cloning and analysis of a cDNA encoding a putative RNA-binding protein, specifically expressed in the rat brain and conserved from Drosophila melanogaster to man. The protein, that contains two regions with chemical homology to double-stranded RNA-binding motifs (16) was called PIPPin after the first four amino acids of the second of these motifs (PIPP, in one-letter code).Here we report that PIPPin c...

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