Neuroscience Letters 381 (2005) 169–174 Minor expression of fascin-1 gene (FSCN1) in NTera2 cells depleted of CREB-binding protein Francesca Megiorni, Paola Indovina, Barbara Mora, Maria Cristina Mazzilli ∗ Department of Experimental Medicine and Pathology, “La Sapienza” University, Viale Regina Elena 324, 00161 Rome, Italy Received 10 December 2004; received in revised form 7 February 2005; accepted 9 February 2005 Abstract CREB-binding protein (CBP) is a transcriptional coactivator whose mutations may cause a generalized perturbation of gene expression. We silenced the CBP gene in NT2 neuronal precursor cells by RNA interference. Hybridization experiments on 1.2 K human cDNA microarrays showed that the FSCN1 gene, encoding for fascin-1 protein, was clearly less expressed in CBP-depleted cells than in controls. This reduction was confirmed by Real Time PCR and Western blotting assays. We also analyzed FSCN1 expression profile during NT2 neuronal differentiation induced by retinoic acid (RA), showing that FSCN1 was up-regulated during neurogenesis. This mRNA increasing suggests the importance of fascin-1 in the formation of mature neurons, in accordance with its actin-bundling function and its localization in neurites and neuronal growth cones. The lower amount of FSCN1 transcript in the absence of the CBP factor was also established in RA-treated cells. In conclusion, this research supports FSCN1 as a novel marker of NT2 neuronal differentiation and the possible role of CBP in its regulation. © 2005 Elsevier Ireland Ltd. All rights reserved. Keywords: CBP transcriptional coactivator; NT2 cells; Small interfering RNA; Microarray analysis; Real Time PCR; Fascin-1 protein CREB-binding protein (CBP) is a coactivator that promotes gene transcription by providing a scaffold for numerous DNA-binding transcription factors and the basic transcrip- tion machinery. Furthermore, CBP contains an intrinsic his- tone acetyltransferase (HAT) activity and may modulate gene expression through acetylation of nucleosomal proteins and various regulatory factors [10,13]. CBP and the closely re- lated p300 protein are involved in multiple physiological processes and several studies indicate that they have sim- ilar functions: chromosomal translocations affecting p300 and CBP genes cause hematological malignancies, and point- mutations or heterozygosity of these genes have been found in various kinds of tumors [13,28]. Despite their extensive similarities, CBP and p300 have unique and different func- tions. Retinoic acid-induced differentiation of F9 embryonal carcinoma cells was inhibited by a ribozyme directed against the p300 mRNA but not by a CBP ribozyme [15]. In a murine model, a full dose of CBP is crucial for hematopoietic stem cell self-renewal; on the contrary, only p300 is essen- ∗ Corresponding author. Tel.: +39 064451286; fax: +39 064454820. E-mail address: cristina.mazzilli@uniromal.it (M.C. Mazzilli). tial for a correct hematopoietic differentiation [25]. In addi- tion, CBP haplo-insufficience is found in Rubinstein–Taybi syndrome (RTS) that is characterized by growth and mental retardation, multiple congenital malformations and increased tumor risk [6,22]. No mutations of the p300 cofac- tor have been described in human congenital diseases. The presence of a particular spectrum of affected tissues in RTS indicates that CBP mutations modify the expression of a re- stricted class of genes, many of which are as yet unidenti- fied. In this work, we propose an in vitro cellular system to dis- sect the biological CBP function in human NTera2/cloneD1 (NT2) neuronal precursor cells [18,23]. In particular, we sup- pressed CBP expression by RNA interference (RNAi) using a specific small interfering RNA (siRNA), and subsequently we looked for genes that presented modified transcriptional pro- files. Microarray, Real Time PCR and Western blotting assays revealed a significant down-expression of the gene encoding the actin-bundling protein fascin-1 (FSCN1) in CBP-depleted cells. Furthermore, we excluded that the reduced amount of FSCN1 gene was attributable to an unspecific effect of type 1 interferon (IFN) activation. 0304-3940/$ – see front matter © 2005 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.neulet.2005.02.027