Short interspersed elements (SINEs) are highly abundant components of mammalian genomes that are propagated by retrotransposition. SINEs are recognized as a causal agent of human disease and must also have had a profound influence in shaping eukaryotic genomes. The B2 SINE family constitutes approximately 0.7% of total mouse genomic DNA (ref. 2) and is also found at low abundance in humans. It resembles the Alu family in several respects, such as its mechanism of propagation. B2 SINEs are derived from tRNA and are transcribed by RNA polymerase (pol) III to generate short transcripts that are not translated. We find here, however, that one B2 SINE also carries an active pol II promoter located outside the tRNA region. Indeed, a B2 element is responsible for the production of a mouse Lama3 transcript. The B2 pol II promoters can be bound and stimulated by the transcription factor USF (for upstream stimulatory factor), as shown by transient transfection experiments. Moreover, this pol II activity does not preclude the pol III transcription necessary for retrotransposition. Dispersal of B2 SINEs by retrotransposition may therefore have provided numerous opportunities for creating regulated pol II transcription at novel genomic sites. This mechanism may have allowed the evolution of new transcription units and new genes.
Several lines of evidence suggest a role for laminin-5 in skin wound healing. We report here that transforming growth factor- (TGF-), which elicits various responses during cutaneous healing, stimulates transcription of the mouse laminin ␣3A (lama3A) gene. To identify the TGF--responsive elements (TGF-REs) on the lama3A promoter, we have generated a series of 5-deletions of the promoter upstream of the -galactosidase reporter gene. Transient cell transfection assays using mouse PAM212 keratinocytes revealed that TGF-REs lie between nucleotides ؊297 and ؊54 relative to the transcription start site. Insertion of the TGF-RE in front of the unresponsive minimal SV40 promoter conferred TGF- inducibility. Computer analysis of the promoter sequence identified three canonical activator protein-1 (AP-1) sites located at nucleotides ؊277 (AP-1A), ؊125 (AP-1B), and ؊69 (AP-1C). Site-directed mutagenesis of either the AP-1A or AP-1C site did not drastically alter the basal activity of the lama3A promoter, but reduced TGF- responsiveness by 50%. Simultaneous mutation of these two AP-1 sites resulted in a 65% decline in the response to TGF-, suggesting a cooperative contribution of each site to the overall promoter activity. In contrast, mutation of the AP-1B site markedly reduced the basal activity of the lama3A promoter, indicating that this AP-1 site is essential for gene expression. Mobility shift assays demonstrated specific binding of Fra-2 and JunD to the AP-1 sites, suggesting for the first time a possible regulatory function for the Fra-2⅐JunD AP-1 complex in a basal keratinocyte-specific gene.Laminin-5 is the major adhesion ligand present in the basement membranes of stratified squamous epithelia (1, 2). In the skin, this adhesive protein is secreted by the basal keratinocytes and colocalize with the anchoring filaments of the lamina lucida of the dermal epidermal junction (3-5). Laminin-5 binds to integrin ␣ 3  1 in focal adhesions and interacts with hemidesmosomes via ␣ 6  4 to form a stable anchorage complex (6, 7). Laminin-5 is a heterotrimeric glycoprotein composed of the ␣3A, 3, and ␥2 polypeptide chains that are products of different genes. Mutations in the genes encoding laminin ␣3 (LA-MA3), 3 (LAMB3), and ␥2 (LAMC2) have been shown to underlie the Herlitz or non-Herlitz forms of junctional epidermolysis bullosa, characterized by blister formation and erosions of the skin and mucosas that frequently lead to neonatal death (8 -13).Several lines of evidence suggest a role for laminin-5 in the re-epithelialization of wound skin repair. Laminin-5 is found at the epidermal-dermal junction at sites and times that coincide with actively migrating or rapidly proliferating basal keratinocytes (14, 15). Moreover, enhancement of laminin-5 transcription is observed at low cell densities in vitro in migrating and proliferating keratinocytes, similar to what happens at the wound edge (15). Although the function of laminin-5 in wound healing remains to be clarified, its major dual contribution would be to all...
Predictive biomarkers for advanced prostate cancer (PCa) are still missing. The sirtuin 7 (SIRT7) has been linked to tumorogenesis but its role in prostate cancer is poorly documented. To determine if SIRT7 can be a biomarker for aggressive prostate cancer and plays a role in PCa aggressiveness. We analyzed the expression of SIRT7 by immunohistochemistry in 57 patients comparing healthy with adjacent cancer tissue. SIRT7 levels were significantly elevated in tumors and its expression was positively associated with the grade. We also demonstrated that the knock down of SIRT7 decreased the migration of DU145 and PC3 cells (two androgen-independent prostate cancer cell lines) whereas the overexpression of the native protein but not the mutated form increased the cell migration and the invasion of the poorly aggressive prostate cancer cell line LNCaP. Finally, we also showed that SIRT7 overexpression induced the resistance to docetaxel. Our results demonstrate that SIRT7 promotes prostate cancer cell aggressiveness and chemoresistance and suggest that SIRT7 is a good predictive biomarker of PCa aggressiveness.
The deregulation of lipid metabolism is a hallmark of tumor cells, and elevated lipogenesis has been reported in prostate cancer. Metformin, a drug commonly prescribed for type II diabetes, displays antitumor properties. Here, we show that metformin inhibits lipogenesis in several prostate cancer cell lines. In LNCaP cells, this effect parallels the decrease of key lipogenic proteins: ACC (acetyl-CoA carboxylase), FASN (fatty acid synthase) and SREBP1c (sterol regulatory element binding protein-1c), whereas there is no modification in DU145 and PC3 cells. Despite the relatively high level of lipogenic proteins induced by the overexpression of a constitutively active form of SREBP1c or treatment with androgens, metformin is still able to inhibit lipogenesis. Metformin does not alter the concentration of malonyl-CoA (the fatty acid precursor), and it only slightly decreases the NADPH levels, which is a co-factor required for lipogenesis, in LNCaP. Finally, we show that the inhibitory effect of metformin on lipogenesis is primarily due to a cellular energy deficit. Metformin decreases ATP in a dose-dependent manner, and this diminution is significantly correlated with the inhibition of lipogenesis in LNCaP and DU145. Indeed, the effect of metformin is linked to changes in the ATP content rather than the regulation of protein expression. Our results describe a new mechanism of action for metformin on prostate cancer metabolism.
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