Chemical Inhibition of Histone Deacetylases 1 and 2 Induces Fetal Hemoglobin through Activation of GATA2.

Therapeutic intervention aimed at reactivation of fetal hemoglobin protein (HbF) is a promising approach for ameliorating sickle cell disease (SCD) and ß-thalassemia. Previous studies showed genetic knockdown of histone deacetylase (HDAC) 1 or 2 is sufficient to induce HbF. Here we show that ACY-957, a selective chemical inhibitor of HDAC1 and 2 (HDAC1/2), elicits a dose and time dependent induction of γ-globin mRNA (HBG) and HbF in cultured primary cells derived from healthy individuals and sickle cell patients. Gene expression profiling of erythroid progenitors treated with ACY-957 identified global changes in gene expression that were significantly enriched in genes previously shown to be affected by HDAC1 or 2 knockdown. These genes included GATA2, which was induced greater than 3-fold. Lentiviral overexpression of GATA2 in primary erythroid progenitors increased HBG, and reduced adult ß-globin mRNA (HBB). Furthermore, knockdown of GATA2 attenuated HBG induction by ACY-957. Chromatin immunoprecipitation and sequencing (ChIP-Seq) of primary erythroid progenitors demonstrated that HDAC1 and 2 occupancy was highly correlated throughout the GATA2 locus and that HDAC1/2 inhibition led to elevated histone acetylation at well-known GATA2 autoregulatory regions. The GATA2 protein itself also showed increased binding at these regions in response to ACY-957 treatment. These data show that chemical inhibition of HDAC1/2 induces HBG and suggest that this effect is mediated, at least in part, by histone acetylation-induced activation of the GATA2 gene.

The androgen receptor cistrome is extensively reprogrammed in human prostate tumorigenesis.

Master transcription factors interact with DNA to establish cell type identity and to regulate gene expression in mammalian cells. The genome-wide map of these transcription factor binding sites has been termed the cistrome. Here we show that the androgen receptor (AR) cistrome undergoes extensive reprogramming during prostate epithelial transformation in man. Using human prostate tissue, we observed a core set of AR binding sites that are consistently reprogrammed in tumors. FOXA1 and HOXB13 colocalized at the reprogrammed AR binding sites in human tumor tissue. Introduction of FOXA1 and HOXB13 into an immortalized prostate cell line reprogrammed the AR cistrome to resemble that of a prostate tumor, functionally linking these specific factors to AR cistrome reprogramming. These findings offer mechanistic insights into a key set of events that drive normal prostate epithelium toward transformation and establish the centrality of epigenetic reprogramming in human prostate tumorigenesis.

Effect of streptozotocin-induced type 1 diabetes mellitus on contraction, calcium transient, and cation contents in the isolated rat heart.

Cations play major physiological and biochemical roles in the excitation-contraction coupling processes in the heart. This study investigated the effect of streptozotocin (STZ)-induced type I diabetes mellitus (DM) on contraction, calcium transient [Ca2+]i, and cation contents in the isolated rat heart compared to age-matched control. Diabetes rats weighed significantly (P < 0.05) less compared to control. They also had significantly (P < 0.05) elevated blood glucose compared to control. The whole heart, as well as the atria, right and left ventricles of the diabetic heart weighed significantly (P < 0.05) less compared to hearts from age control rats. The force of contraction and time to peak (t-pk) contraction in diabetic ventricular myocytes increased significantly (P < 0.05) compared to control. By contrast, these parameters did not change for the Ca2+ transient except for the time to half (t(1/2)) relaxation. The levels of sodium (Na+), potassium (K+), calcium (Ca2+), magnesium (Mg2+), iron (Fe2+), copper (Cu2+), and zinc (Zn2+) in the hearts varied from diabetic compared to control animals. The results indicate that 6-8 weeks of STZ-induced DM is associated with marked changes in contraction and in cation contents of the heart. The delay in the t(1/2) relaxation of the Ca2+ transient may be responsible for the elevated contraction seen in the diabetic heart. Moreover, the changes in cation contents in the heart may be responsible for abnormal cardiac rhythms and activity during DM.

Contraction and cation contents of skeletal soleus and EDL muscles in age-matched control and diabetic rats.

Skeletal muscle atrophy and neuropathy are two of the major long-term complications of diabetes mellitus (DM). This article investigates the effect of streptozotocin (STZ)-induced type 1 DM on contraction and cation contents in soleus and EDL muscles compared to age-matched control rats. Adult Wistar rats were humanely killed and the soleus and EDL muscles located and excised rapidly and placed in organ baths. The muscles were electrically stimulated (EFS, 50 V, 1 ms, 1-50 Hz) and isometric contraction monitored using a chart recorder. At the end of the experiments the muscles were blotted, weighed, and dissolved in concentrated nitric acid for the measurements of cation contents. Diabetic rats have significantly (P < 0.05) reduced body and muscle weights and significantly (P < 0.05) elevated blood glucose (29.65 +/- 0.33 mM, n = 10) compared to control (6.06 +/- 0.21 mM, n = 10). EFS evoked frequency dependent contraction in both soleus and EDL muscles of healthy control rats. The response was significantly (P < 0.05, n = 8) increased in EDL compared to soleus. In the diabetic soleus and EDL muscles the force of contraction was significantly (P < 0.05, n = 8) reduced in both muscle types compared to control. The levels of total sodium (Na+), potassium (K+), calcium (Ca2+), magnesium (Mg2+), zinc (Zn2+), copper (Cu2+), and iron (Fe2+) were significantly (P < 0.05) reduced in diabetic soleus and EDL muscles compared to control. The results indicate that STZ-induced skeletal muscle atrophy is associated with marked reductions in both the force of contraction and cation contents in EDL and soleus muscles of the rat.

Effects of streptozotocin-induced type 1 diabetes mellitus on total protein concentrations and cation contents in the isolated pancreas, parotid, submandibular, and lacrimal glands of rats.

This study investigated the effect of streptozotocin (STZ)-induced type 1 diabetes mellitus (DM) on total protein concentration and levels of sodium (Na+), potassium (K+), magnesium (Mg2+), zinc (Zn2+), copper (Cu2+), calcium (Ca2+), and iron (Fe2+) in the pancreas, parotid, submandibular, and lacrimal glands of the rat, compared to age-matched control animals. Protein concentrations were measured by the Bradford Assay, whereas levels of Na+, K+, Mg2+, Zn2+, Cu2+, Ca2+, and Fe2+ were measured by flame photometry and atomic absorbance spectrophotometry. The results show marked changes in the characteristics of diabetic and control animals. Diabetic rats and their different glands weighed significantly (P < 0.05) less compared to age-matched controls. Diabetic rats also have significantly elevated blood glucose and significantly reduced plasma insulin, compared to controls. The results also show that the concentrations of proteins and levels of cations were significantly (P < 0.05) reduced in the pancreas, parotid, submandibular and lacrimal glands of diabetic rats, compared to glands from age-matched animals. These differences in the cation contents and protein levels in STZ-induced DM in this study, along with supporting evidences from previous studies, may provide evidence for the development of long-term complications of DM including exocrine gland deficiencies.