Initial adhesion suppression of biofilm-forming and copper-tolerant bacterium Variovorax sp. on laser microtextured copper surfaces.

The growth of detrimental biofilms on metal surfaces affects their structural performance and lifespan. Microtopographic texturization has emerged as an approach to suppress biofilm growth by preventing the initial stages of bacterial adhesion. This work studies the effects of linear pattern copper texturization on the initial adhesion steps of the biofilm-forming and copper-resistant bacterium Variovorax sp. Linear patterns with 4.7, 6.8, 14, and 18 µm periodicity were produced by direct laser interference patterning (DLIP) on copper coupons. Surface features were characterized by microscopic and spectroscopic techniques, and bacterial adhesion behavior was characterized by epifluorescence microscopy and functionalization of atomic force microscopy tips. We found a periodicity of 4.7 µm as the most efficient pattern to suppress Variovorax sp. initial adhesion by 31.1 % with respect to the nontextured surface. Preferential settlement in hummocks over hollows was observed for patterns with 14 and 18 µm periodicity, with adhesion events showing higher frequency in these topographies than patterns with periodicities of 4.7 and 6.8 µm. Our results highlight an understanding of the initial bacteria-copper adhesion and settlement behavior, thus contributing to the potential development of innocuous strategies for controlling biofilm growth on copper-based materials.

Targeting the CoREST complex with dual histone deacetylase and demethylase inhibitors.

Here we report corin, a synthetic hybrid agent derived from the class I HDAC inhibitor (entinostat) and an LSD1 inhibitor (tranylcypromine analog). Enzymologic analysis reveals that corin potently targets the CoREST complex and shows more sustained inhibition of CoREST complex HDAC activity compared with entinostat. Cell-based experiments demonstrate that corin exhibits a superior anti-proliferative profile against several melanoma lines and cutaneous squamous cell carcinoma lines compared to its parent monofunctional inhibitors but is less toxic to melanocytes and keratinocytes. CoREST knockdown, gene expression, and ChIP studies suggest that corin's favorable pharmacologic effects may rely on an intact CoREST complex. Corin was also effective in slowing tumor growth in a melanoma mouse xenograft model. These studies highlight the promise of a new class of two-pronged hybrid agents that may show preferential targeting of particular epigenetic regulatory complexes and offer unique therapeutic opportunities.

Characterizing HSF1 Binding and Post-Translational Modifications of hsp70 Promoter in Cultured Cortical Neurons: Implications in the Heat-Shock Response.

Causes of lower induction of Hsp70 in neurons during heat shock are still a matter of debate. To further inquire into the mechanisms regulating Hsp70 expression in neurons, we studied the activity of Heat Shock Factor 1 (HSF1) and histone posttranslational modifications (PTMs) at the hsp70 promoter in rat cortical neurons. Heat shock induced a transient and efficient translocation of HSF1 to neuronal nuclei. However, no binding of HSF1 at the hsp70 promoter was detected while it bound to the hsp25 promoter in cortical neurons during heat shock. Histone PTMs analysis showed that the hsp70 promoter harbors lower levels of histone H3 and H4 acetylation in cortical neurons compared to PC12 cells under basal conditions. Transcriptomic profiling data analysis showed a predominant usage of cryptic transcriptional start sites at hsp70 gene in the rat cerebral cortex, compared with the whole brain. These data support a weaker activation of hsp70 canonical promoter. Heat shock increased H3Ac at the hsp70 promoter in PC12 cells, which correlated with increased Hsp70 expression while no modifications occurred at the hsp70 promoter in cortical neurons. Increased histone H3 acetylation by Trichostatin A led to hsp70 mRNA and protein induction in cortical neurons. In conclusion, we found that two independent mechanisms maintain a lower induction of Hsp70 in cortical neurons. First, HSF1 fails to bind specifically to the hsp70 promoter in cortical neurons during heat shock and, second, the hsp70 promoter is less accessible in neurons compared to non-neuronal cells due to histone deacetylases repression.

Differential properties of transcriptional complexes formed by the CoREST family.

Mammalian genomes harbor three CoREST genes. rcor1 encodes CoREST (CoREST1), and the paralogues rcor2 and rcor3 encode CoREST2 and CoREST3, respectively. Here, we describe specific properties of transcriptional complexes formed by CoREST proteins with the histone demethylase LSD1/KDM1A and histone deacetylases 1 and 2 (HDAC1/2) and the finding that all three CoRESTs are expressed in the adult rat brain. CoRESTs interact equally strongly with LSD1/KDM1A. Structural analysis shows that the overall conformation of CoREST3 is similar to that of CoREST1 complexed with LSD1/KDM1A. Nonetheless, transcriptional repressive capacity of CoREST3 is lower than that of CoREST1, which correlates with the observation that CoREST3 leads to a reduced LSD1/KDM1A catalytic efficiency. Also, CoREST2 shows a lower transcriptional repression than CoREST1, which is resistant to HDAC inhibitors. CoREST2 displays lower interaction with HDAC1/2, which is barely present in LSD1/KDM1A-CoREST2 complexes. A nonconserved leucine in the first SANT domain of CoREST2 severely weakens its association with HDAC1/2. Furthermore, CoREST2 mutants with increased HDAC1/2 interaction and those without HDAC1/2 interaction exhibit equivalent transcriptional repression capacities, indicating that CoREST2 represses in an HDAC-independent manner. In conclusion, differences among CoREST proteins are instrumental in the modulation of protein-protein interactions and catalytic activities of LSD1/KDM1A-CoREST-HDAC complexes, fine-tuning gene expression regulation.

CoREST represses the heat shock response mediated by HSF1.

The stress response in cells involves a rapid and transient transcriptional activation of stress genes. It has been shown that Hsp70 limits its own transcriptional activation functioning as a corepressor of heat shock factor 1 (HSF1) during the attenuation of the stress response. Here we show that the transcriptional corepressor CoREST interacts with Hsp70. Through this interaction, CoREST represses both HSF1-dependent and heat shock-dependent transcriptional activation of the hsp70 promoter. In cells expressing short hairpin RNAs directed against CoREST, Hsp70 cannot repress HSF1-dependent transcription. A reduction of CoREST levels also provoked a significant increase of Hsp70 protein levels and an increase of HSF1-dependent transactivation of hsp70 promoter. Via chromatin immunoprecipitation assays we show that CoREST is bound to the hsp70 gene promoter under basal conditions and that its binding increases during heat shock response. In conclusion, we demonstrated that CoREST is a key regulator of the heat shock stress response.