Histone deacetylases (HDACs) form a family of enzymes, which have fundamental roles in the epigenetic regulation of gene expression and contribute to the growth, differentiation, and apoptosis of cancer cells. In this study, we further investigated the biological function of HDAC5 in cancer cells. We found HDAC5 is associated with actively replicating pericentric heterochromatin during late S phase. We demonstrated that specific depletion of HDAC5 by RNA interference resulted in profound changes in the heterochromatin structure and slowed down ongoing replication forks. This defect in heterochromatin maintenance and assembly are sensed by DNA damage checkpoint pathways, which triggered cancer cells to autophagy and apoptosis, and arrested their growth both in vitro and in vivo. Finally, we also demonstrated that HDAC5 depletion led to enhanced sensitivity of DNA to DNAdamaging agents, suggesting that heterochromatin de-condensation induced by histone HDAC5 silencing may enhance the efficacy of cytotoxic agents that act by targeting DNA in vitro. Together, these results highlighted for the first time an unrecognized link between HDAC5 and the maintenance/assembly of heterochromatin structure, and demonstrated that its specific inhibition might contribute to increase the efficacy of DNA alteration-based cancer therapies in clinic.

The rapidity and the simplicity of this approach could represent a suitable alternative to the other protocols. Additionally, this approach could be favourable for high- throughput applications in intracellular and intratissular metabolite measurements.

Histone deacetylases (HDACs) are a family of 18 enzymes that deacetylate lysine residues of both histone and nonhistone proteins and to a large extent govern the process of angiogenesis. Previous studies have shown that specific inhibition of HDAC7 blocks angiogenesis both in vitro and in vivo. However, the underlying molecular mechanisms are not fully understood and hence preclude any meaningful development of suitable therapeutic modalities. The goal of the present study was to further the understanding of HDAC7 epigenetic control of angiogenesis in human endothelial cells using the proteomic approach. The underlying problem was approached through siRNA-mediated gene-expression silencing of HDAC7 in human umbilical vein endothelial cells (HUVECs). To this end, HUVEC proteins were extracted and proteomically analyzed. The emphasis was placed on up-regulated proteins, as these may represent potential direct epigenetic targets of HDAC7. Among several proteins, A-kinase anchor protein 12 (AKAP12) was the most reproducibly up-regulated protein following HDAC7 depletion. This overexpression of AKAP12 was responsible for the inhibition of migration and tube formation in HDAC7-depleted HUVEC. Mechanistically, H3 histones associated with AKAP12 promoter were acetylated following the removal of HDAC7, leading to an increase in its mRNA and protein levels. AKAP12 is responsible for protein kinase C mediated phosphorylation of signal transducer and activator of transcription 3 (STAT3). Phosphorylated STAT3 increasingly binds to the chromatin and AKAP12 promoter and is necessary for maintaining the elevated levels of AKAP12 following HDAC7 knockdown. We demonstrated for the first time that AKAP12 tumor/angiogenesis suppressor gene is an epigenetic target of HDAC7, whose elevated levels lead to a negative regulation of HUVEC migration and inhibit formation of tube-like structures.

The US FDA approval of broad-spectrum histone deacetylase (HDAC) inhibitors has firmly laid the cancer community to explore HDAC inhibition as a therapeutic approach for cancer treatment. Hitting one HDAC member could yield clinical benefit but this required a complete understanding of the functions of the different HDAC members. Here we explored the consequences of specific HDAC5 inhibition in cancer cells. We demonstrated that HDAC5 inhibition induces an iron-dependent reactive oxygen species (ROS) production, ultimately leading to apoptotic cell death as well as mechanisms of mitochondria quality control (mitophagy and mitobiogenesis). Interestingly, adaptation of HDAC5-depleted cells to oxidative stress passes through reprogramming of metabolic pathways towards glucose and glutamine. Therefore, interference with both glucose and glutamine supply in HDAC5-inhibited cancer cells significantly increases apoptotic cell death and reduces tumour growth in vivo; providing insight into a valuable clinical strategy combining the selective inhibition of HDAC5 with various inhibitors of metabolism as a new therapy to kill cancer cells.

Telomeres are major regulators of genome stability and cell proliferation. A detailed understanding of the mechanisms involved in their maintenance is of foremost importance. Of those, telomere chromatin remodeling is probably the least studied; thus, we intended to explore the role of a specific histone deacetylase on telomere maintenance. We uncovered a new role for histone deacetylase 5 (HDAC5) in telomere biology. We report that HDAC5 is recruited to the long telomeres of osteosarcoma- and fibrosarcoma-derived cell lines, where it ensures proper maintenance of these repetitive regions. Indeed, depletion of HDAC5 by RNAi resulted in the shortening of longer telomeres and homogenization of telomere length in cells that use either telomerase or an alternative mechanism of telomere maintenance. Furthermore, we present evidence for the activation of telomere recombination on depletion of HDAC5 in fibrosarcoma telomerase-positive cancer cells. Of potential importance, we also found that depletion of HDAC5 sensitizes cancer cells with long telomeres to chemotherapeutic drugs. Cells with shorter telomeres were used to control the specificity of HDAC5 role in the maintenance of long telomeres. HDAC5 is essential for the length maintenance of long telomeres and its depletion is required for sensitization of cancer cells with long telomeres to chemotherapy.