BRPF3 knockdown or inhibition moderately reverses olaparib resistance in high grade serous ovarian carcinoma, but depletion of H3K14 acetylation has no effect

Background: PARP inhibitors (PARPi) kill cancer cells by stalling DNA replication and preventing DNA repair, resulting in a critical accumulation of DNA damage. Resistance to PARPi is a growing clinical problem in the treatment of high grade serous ovarian carcinoma (HGSOC). Acetylation of histone H3 lysine 14 (H3K14ac) and associated histone acetyltransferases (HATs) have known functions in DNA repair and replication, but their expression and activities have not been examined in the context of PARPi-resistant HGSOC. Results: Using mass spectrometry profiling of histone modifications, we observed altered H3K14ac enrichment in PARPi-resistant HGSOC cells relative to isogenic PARPi-sensitive lines. By RT-qPCR and RNA-Seq, we also observed altered expression of numerous HATs in PARPi-resistant HGSOC cells and a PARPi-resistant PDX model. Knockdown of HATs only modestly altered PARPi response, although knockdown and inhibition of PCAF significantly increased resistance. Pharmacologic inhibition of HBO1 severely depleted H3K14ac but did not affect PARPi response. However, knockdown and inhibition of BRPF3, which is known to interact in a complex with HBO1, did reduce PARPi resistance. Conclusions: This study demonstrates that severe depletion of H3K14ac does not affect PARPi response in HGSOC. Our data suggest that bromodomain functions of HAT proteins such as PCAF, or accessory proteins such as BRPF3, may play a greater role in PARPi response than acetyltransferase functions.

IL-1β and TNFα Cooperativity in Regulating IL-6 Expression in Adipocytes Depends on CREB Binding and H3K14 Acetylation

IL-6 was found to be overexpressed in the adipose tissue of obese individuals, which may cause insulin resistance. However, the regulation of IL-6 in adipocytes in obesity setting remains to be explored. Since IL-1β and TNFα are increased in obese adipose tissue and promote inflammation, we investigated whether cooperation between IL-1β and TNFα influences the production of IL-6. Our data show that IL-1β and TNFα cooperatively enhances IL-6 expression in 3T3L-1 adipocytes. Similar results were seen in human adipocytes isolated from subcutaneous and visceral fat. Although adipocytes isolated from lean and obese adipose tissues showed similar responses for production of IL-6 when incubated with IL-1β/TNFα, secretion of IL-6 was higher in adipocytes from obese tissue. TNFα treatment enhanced CREB binding at CRE locus, which was further enhanced with IL-1β, and was associated with elevated histone acetylation at CRE locus. On the other hand, IL-1β treatments mediated C/EBPβ binding to NF-IL-6 consensus, but not sufficiently to mediate significant histone acetylation. Interestingly, treatment with both stimulatory factors amplifies CREB binding and H3K14 acetylation. Furthermore, histone acetylation inhibition by anacardic acid or curcumin reduces IL-6 production. Notably, inhibition of histone deacetylase (HDAC) activity by trichostatin A (TSA) resulted in the further elevation of IL-6 expression in response to combined treatment of adipocytes with IL-1β and TNFα. In conclusion, our results show that there is an additive interaction between IL-1β and TNFα that depends on CREB binding and H3K14 acetylation, and leads to the elevation of IL-6 expression in adipocytes, providing interesting pathophysiological connection among IL-1β, TNFα, and IL-6 in settings such as obesity.

Bromodomain-containing protein BRPF1 is a therapeutic target for liver cancer

Epigenetic deregulation plays an essential role in hepatocellular carcinoma (HCC) progression. Bromodomains are epigenetic “readers” of histone acetylation. Recently, bromodomain inhibitors have exhibited promising therapeutic potential for cancer treatment. Using transcriptome sequencing, we identified BRPF1 (bromodomain and PHD finger containing 1) as the most significantly upregulated gene among the 43 bromodomain-containing genes in human HCC. BRPF1 upregulation was significantly associated with poor patient survival. Gene ablation or pharmacological inactivation of BRPF1 significantly attenuated HCC cell growth in vitro and in vivo. BRPF1 was involved in cell cycle progression, senescence and cancer stemness. Transcriptome sequencing revealed that BRPF1 is a master regulator controlling the expression of multiple key oncogenes, including E2F2 and EZH2. We demonstrated that BRPF1 activated E2F2 and EZH2 expression by facilitating promoter H3K14 acetylation through MOZ/MORF complex. In conclusion, BRPF1 is frequently upregulated in human HCCs. Targeting BRPF1 may be an approach for HCC treatment.

The Transcriptional Adaptor Protein ADA3a Modulates Flowering of Arabidopsis thaliana

Histone acetylation is directly related to gene expression. In yeast, the acetyltransferase general control nonderepressible-5 (GCN5) targets histone H3 and associates with transcriptional co-activators alteration/deficiency in activation-2 (ADA2) and alteration/deficiency in activation-3 (ADA3) in complexes like SAGA. Arabidopsis thaliana has two genes encoding proteins, designated ADA3a and ADA3b, that correspond to yeast ADA3. We investigated the role of ADA3a and ADA3b in regulating gene expression during flowering time. Specifically, we found that knock out mutants ada3a-2 and the double mutant ada3a-2 ada3b-2 lead to early flowering compared to the wild type plants under long day (LD) conditions and after moving plants from short days to LD. Consistent with ADA3a being a repressor of floral initiation, FLOWERING LOCUS T (FT) expression was increased in ada3a mutants. In contrast, other genes involved in multiple pathways leading to floral transition, including FT repressors, players in GA signaling, and members of the SPL transcriptional factors, displayed reduced expression. Chromatin immunoprecipitation analysis revealed that ADA3a affects the histone H3K14 acetylation levels in SPL3, SPL5, RGA, GAI, and SMZ loci. In conclusion, ADA3a is involved in floral induction through a GCN5-containing complex that acetylates histone H3 in the chromatin of flowering related genes.

HDAC3 maintains oocyte meiosis arrest by repressing amphiregulin expression before the LH surge

It is known that granulosa cells (GCs) mediate gonadotropin-induced oocyte meiosis resumption by releasing EGF-like factors in mammals, however, the detailed molecular mechanisms remain unclear. Here, we demonstrate that luteinizing hormone (LH) surge-induced histone deacetylase 3 (HDAC3) downregulation in GCs is essential for oocyte maturation. Before the LH surge, HDAC3 is highly expressed in GCs. Transcription factors, such as FOXO1, mediate recruitment of HDAC3 to the amphiregulin (Areg) promoter, which suppresses AREG expression. With the LH surge, decreased HDAC3 in GCs enables histone H3K14 acetylation and binding of the SP1 transcription factor to the Areg promoter to initiate AREG transcription and oocyte maturation. Conditional knockout of Hdac3 in granulosa cells in vivo or inhibition of HDAC3 activity in vitro promotes the maturation of oocytes independent of LH. Taking together, HDAC3 in GCs within ovarian follicles acts as a negative regulator of EGF-like growth factor expression before the LH surge.